Methods for the characterization of fluid

Abstract

Provided herein are methods for detecting a menstrual cycle disorder. Menstrual cycle disorders is detected through analysis of a property of a collected menstrual fluid sample, such as gene expression, flow rate, protein content, nucleic acid content, biomarkers, or other properties.

Description

CROSS-REFERENCE

[0001] This application claims the benefit of U.S. Patent Application No. 62 / 929,579, filed November 1, 2019, U.S. Patent Application No. 62 / 930,465, filed November 4, 2019, and U.S. Patent Application No. 63 / 061,709, filed August 5, 2020, which are hereby incorporated by reference in their entirety. BACKGROUND

[0002] Women are underrepresented in research. Diagnostic development for women’s reproductive health has been hampered by a lack of understanding of uterine and menstrual physiology. To date, none of the peripheral blood biomarkers proposed for diagnosing women’s reproductive health exhibit the accuracy required for clinical use.

[0003] Not only have period movements allowed for reduced stigma and national conversations about menstruation, but women are also gravitating to consumer health and digital recruitment platforms, creating new opportunities for research and diagnostic development. Menstrual blood provides direct access to reproductive tissue. SUMMARY

[0004] In certain aspects, disclosed herein are methods of using a predictive model to identify a condition of heavy menstrual bleeding (HMB). In some embodiments, the method comprises, using one or more processors in a computer server: (a) receiving data defining a plurality of subject digital biomarkers, each digital biomarker comprising a response by the subject to an associated inquiry in a computing data storage; (b) retrieving from computer data storage a combination of independent variables relating to the subject; (c) using a predictive model to determine a dependent variable representing a subject HMB risk score for the subject based on the combination of independent variables relating to the subject; (d) determining if the dependent variable representing the subject HMB score is above a threshold; and if the dependent variable representing the subject HMB score is above the threshold, sending information to be displayed. In some embodiments, the combination of independent variables includes objective data relating to the subject’s menstrual bleeding. In some embodiments, the combination of independent variables includes survey data relating to the subject. In some embodiments, the survey data comprises at least one independent variable selected from the 2020373103   25 Mar 2026 group consisting of a menstrual cycle phenotype, a physical body characteristic, a disease or condition, a medical treatment, demographic information, lifestyle information, ancestry, sexuality, menstrual management, health care usage and access. In some embodiments, the combination of independent variables includes health record data relating to the subject. In some embodiments, the objective data comprises a menstrual flow rate measurement. In some embodiments, the menstrual flow rate measurement comprises: (a) collecting menstrual fluid from a subject for a specified duration; (b) measuring the volume of collected menstrual fluid; (c) calculating a flow rate from the volume and the specified duration. In some embodiments, the method further comprises analyzing a biological marker from the collected menstrual fluid. In some embodiments, the biological marker is selected from a cell-type, a protein, a microorganism, a metabolite, a hematocrit level, or a nucleic acid. In some embodiments, the biological marker is unique to menstrual fluid.

[0005] Described herein, in some embodiments, are methods for generating a severity assessment of a menstrual bleeding state of a human female subject. In some embodiments, the method comprises: (a) presenting one or more questions to the subject about a first set of attributes related to the subject’s menstrual history and a second set of attributes related to a subject’s menstrual phenotype, wherein the one or more questions are presented to the subject on a display of a graphic user interface of an input device; (b) prompting the subject to enter a response to the one or more questions into the input device, wherein the input device transmits the response to a system comprising a processor and a computer-readable memory, wherein the system calculates an assessment score corresponding to menstrual bleeding state of the subject using the response to the one or more questions and stores the assessment score in the memory; (c) using an assay to perform one or more measurements on a menstrual fluid sample from the subject; (d) comparing the one or more measurements with one or more predetermined thresholds; and determine based on the calculated assessment score and the comparison with the one or more predetermined thresholds, the menstrual bleeding state of the subject; and generating a severity assessment of the subject’s menstrual bleeding state.

[0006] Described herein, in certain aspects are methods of preparation of a biological sample. In some embodiments, the method comprises: (a) identifying a subject; (b) classifying the subject into a risk group based on one or more digital biomarkers; (c) collecting menstrual fluid from a subject for a specified duration; (d) measuring the volume of collected menstrual fluid €calculating a flow rate from the volume and the specified duration. In some embodiments, the collecting step is performed using a cup, a tampon, or a pad. In some embodiments, the 2020373103   25 Mar 2026 menstrual fluid is collected in a manner that preserves at least one of intact cells from the vaginal-cervical space, protein, metabolite, DNA or RNA. In some embodiments, the collecting step comprises contacting the menstrual fluid or a portion thereof with a preserving solution. In some embodiments, the method further comprises extracting nucleic acid from the menstrual fluid and measuring at least one nucleic acid parameter. In some embodiments, the method further comprises measuring the presence of level of a metabolite in the menstrual fluid. In some embodiments, the nucleic acid parameter comprises the amount of nucleic acid, the diversity of nucleic acid, the presence of a miRNA, mRNA expression, copy number. In some embodiments, the method further comprises separating or isolating one or more cell types from the menstrual fluid. In some embodiments, the one or more cell types are selected from the group selected from immune cells, ovarian cells, fallopian tube cells, endometrial cells, and cervical cells. In some embodiments, the method further comprises repeating the collecting step for two or more longitudinal samples from the subject. In some embodiments, the method further comprises measuring an individual flow rate for each longitudinal sample and deriving a mean, average or progression of flow rate from the individual flow rates. In some embodiments, the digital biomarker comprises a factor listed in Table 4. Described herein, in certain aspects is a method for detecting a menstrual cycle disorder, comprising: (a) determining an expression level of one or more markers in a fluid sample obtained from the vaginal cavity of a subject, wherein the one or more markers are selected from Table 1, Table 2, and / or Table 3; and (b) comparing said expression level to a reference level of said one or more markers; wherein an increased or decreased expression level of said one or more markers relative to said reference expression level indicates that said subject has said menstrual cycle disorder. In some embodiments, the fluid sample is obtained from the subject during the subject’s menstrual window. In some embodiments, the reference level is obtained from the subject in a time period outside subject’s menstrual window. In some embodiments, the reference level is obtained from a healthy control subject or an average level from a group of healthy control subjects. In some embodiments, the one or more markers are protein expression markers selected from Table 1. In some embodiments, the one or more markers are gene expression markers selected from Table 2. In some embodiments, the one or more markers are gene expression markers selected from Table 3. In some embodiments, said menstrual cycle disorder is heavy menstrual bleeding. In some embodiments, the method further comprises determining a risk level for the menstrual cycle disorder in the subject. In some embodiments, the determining a risk level step comprises assessing one or more phenotypic or behavioral characteristics of the subject selected from 2020373103   25 Mar 2026 Table 4 and / or Table 5. In some embodiments, the method further comprises stratifying the subject into a treatment group based on the risk level. In some embodiments, the method further comprises obtaining two or more fluid samples from the vaginal cavity of the same subject, wherein the two or more fluid samples are from different time points within the subject’s menstrual cycle, wherein step (a) is repeated for each of the two or more fluid samples and wherein the expression level for each of the two or more fluid samples are compared in step (b). In some embodiments, the fluid sample comprises blood. In some embodiments, the fluid sample comprises shed endothelial cells and shed epithelial cells. In some embodiments, the fluid sample comprises a cell type selected from the group consisting of endothelial cells, epithelial cells, immune cells, and stem cells. In some embodiments, the two or more fluid samples are obtained during the subject’s menstrual window. In some embodiments, at least one of the two or more fluid samples are obtained outside of the subject’s menstrual window. In some embodiments, the one or more markers are selected from the group consisting of HLA-Aa (Major histocompatibility complex class I), IL-6ST (Interleukin 6 signal transducer), APCDD1L (Adenomatosis polyposis coli downregulated 1-like ), TBX3 (T-box 3 ), UBN1a (Ubinuclein), DSPa (Desmoplakin) , SDCBP2 (Syndecan binding protein (syntenin)) , EZH2 (Enhancer of zeste 2 polycomb repressive complex 2 subunit) , UHMK1 (U2AF homology motif (UHM) kinase), NR2C2 (Nuclear receptor subfamily 2, group C, member 2), MIR1282a (microRNA), TMED6a (Transmembrane p24 trafficking protein), VAV3 (Vav 3 guanine nucleotide exchange factor), CDC42BPA (CDC42 binding protein kinase alpha (DMPK-like)) , C17orf75 (Chromosome 17 open reading frame), MB21D1 (Mab-21 domain containing 1), PTPRC (Protein tyrosine phosphatase, receptor type C), WISP1 (WNT1 inducible signaling pathway protein 1), CDC27 (Cell division cycle 27), FSD1L (Fibronectin type III and SPRY domain containing 1-like), BPIFB1 ((C20orf114)a BPI fold containing family B, member 1), SCGB3A1a (Secretoglobin, family 3A, member 1) TFF3a (Trefoil factor 3 (intestinal)), SCGB1D2a (Secretoglobin, family 1D, member 2), SCGB2A2a (Secretoglobin, family 2A, member 2), PRODH (Proline dehydrogenase (oxidase) 1), MFF (Mitochondrial fission factor), TSPAN8 (Tetraspanin), CXCL6a (Chemokine (C-X-C motif) ligand 6), SPDYE2 (Speedy / RINGO cell cycle regulator family member E2), FCGBP (Fc fragment of IgG binding protein), IRX6 (Iroquois homeobox 6), ADAM10 (ADAM metallopeptidase domain 10), MUC5ACa (Mucin 5AC, oligomeric mucus / gel-forming), TRHDE-AS1 (TRHDE antisense RNA (LOC283392)), CYP26A1 (Cytochrome P450, family 26, subfamily A, polypeptide 1), SAA2a (Serum amyloid A2), MMEL1 (Membrane metalloendopeptidase-like 1), GRIP2 2020373103   25 Mar 2026 (Glutamate receptor interacting protein 2), and SAA1a (Serum amyloid A1). In some embodiments, the method further comprises determining an increased expression level of one or more markers selected from the group consisting of HLA-Aa (Major histocompatibility complex class I), IL-6ST (Interleukin 6 signal transducer), APCDD1L (Adenomatosis polyposis coli downregulated 1-like ), TBX3 (T-box 3 ), UBN1a (Ubinuclein), DSPa (Desmoplakin) , SDCBP2 (Syndecan binding protein (syntenin)) , EZH2 (Enhancer of zeste 2 polycomb repressive complex 2 subunit) , UHMK1 (U2AF homology motif (UHM) kinase), NR2C2 (Nuclear receptor subfamily 2, group C, member 2), MIR1282a (microRNA), TMED6a (Transmembrane p24 trafficking protein), VAV3 (Vav 3 guanine nucleotide exchange factor), CDC42BPA (CDC42 binding protein kinase alpha (DMPK-like)) , C17orf75 (Chromosome 17 open reading frame), MB21D1 (Mab-21 domain containing 1), PTPRC (Protein tyrosine phosphatase, receptor type C), WISP1 (WNT1 inducible signaling pathway protein 1), CDC27 (Cell division cycle 27), and FSD1L (Fibronectin type III and SPRY domain containing 1-like) relative to said reference expression level. In some embodiments, the method further comprises determining a decreased expression level of the one or more markers selected from the group consisting of BPIFB1 ((C20orf114)a BPI fold containing family B, member 1), SCGB3A1a (Secretoglobin, family 3A, member 1) TFF3a (Trefoil factor 3 (intestinal)), SCGB1D2a (Secretoglobin, family 1D, member 2), SCGB2A2a (Secretoglobin, family 2A, member 2), PRODH (Proline dehydrogenase (oxidase) 1), MFF (Mitochondrial fission factor), TSPAN8 (Tetraspanin), CXCL6a (Chemokine (C-X-C motif) ligand 6), SPDYE2 (Speedy / RINGO cell cycle regulator family member E2), FCGBP (Fc fragment of IgG binding protein), IRX6 (Iroquois homeobox 6), ADAM10 (ADAM metallopeptidase domain 10), MUC5ACa (Mucin 5AC, oligomeric mucus / gel-forming), TRHDE-AS1 (TRHDE antisense RNA (LOC283392)), CYP26A1 (Cytochrome P450, family 26, subfamily A, polypeptide 1), SAA2a (Serum amyloid A2), MMEL1 (Membrane metalloendopeptidase-like 1), GRIP2 (Glutamate receptor interacting protein 2), and SAA1a (Serum amyloid A1).relative to said reference expression level. In some embodiments, the method further comprises determining an expression pattern of said one or more genes or expression products thereof. In some embodiments, the fluid sample is disposed in a sample collector. In some embodiments, said sample collector is a pad, a tampon, a vaginal cup, a cervical cap, a menstrual disk, a cervical disk, a sponge, or an interlabial pad. In some embodiments, the sample collector comprises a chamber comprising a buffer for preserving intact cells, DNA, RNA, protein, metabolite(s), or any combination thereof. In some embodiments, the method further comprises the step of treating the subject for a heavy 2020373103   25 Mar 2026 menstrual bleeding disorder if the increased or decreased expression level of said one or more markers relative to said reference expression level indicates that said subject has said menstrual cycle disorder. In some embodiments, the step of treating is selected from the group consisting of a therapeutic agent, a surgical intervention or a combination thereof. In some embodiments, the therapeutic agent is selected from the group consisting of an antifibrinolytic agent, a combined hormonal contraceptive, a progestogens, a progestogen-releasing intrauterine device, an androgen, a gonadotropin releasing hormone analogue or any combination thereof. In some embodiments, the surgical intervention is selected from the group consisting of surgical excision, n endometrial ablation, endometrial cryoablation, uterine artery embolization, myomectomy, hysterectomy, and any combination thereof. In some embodiments, the method further comprises placing the subject in a non-treatment category if the subject does not have increased or decreased expression level of said one or more markers relative to said reference expression level.

[0007] In certain aspects, disclosed herein are methods of producing a desired preparation for assessment of menstrual health In some embodiments, the method comprises: (a) receiving a fluid sample collected from the vaginal cavity of a subject, the fluid sample comprising one or more types of cells; (b) contacting the fluid sample with a buffer solution under conditions suitable to maintain one or more cell types in a substantially intact state; (c) separating one cell type in the fluid sample from the remaining fluid sample; (d) determining an expression level of one or more markers in the one cell type, wherein the one or more markers are selected from Table 1 and / or Table 2; and (e) comparing the expression level with a reference level to assess a level of menstrual health. In some embodiments, step (b) maintains at least 90%, 95%, or substantially 100% of said one or more types of cells in a substantially intact state. In some embodiments, the fluid sample is a menstrual fluid sample. In some embodiments, the one cell type is selected from the group consisting of endothelial cells, epithelial cells, mesenchymal cells, and leukocytes. In some embodiments, the one cell type is separated based on expression of a cell surface antigen. In some embodiments, the one cell type is an endothelial cell and the cell surface antigen is selected from the group consisting of CD31 / PECAM-1, CD34, CD36 / SR-B3, CD39, CD44, CD47, CD54 / ICAM-1, CD61, CD62E, CD62P, CD80, CD86, CD93, CD102, CD105, CD106, CD112, CD117, ESAM, ENDOMUCIN, CXCL16, CD121a, CD141, CD142, CD143, CD144, CD146, CD147, CD151, CD160, CD201, CD213a, CD248, CD309, ADAMS 8-17, 33, ADAMTS-13, ADAMTS-18, VWF, TEM8, NOTCH, KLF4 and any combination thereof. In some embodiments, the one cell type is an epithelial cell and the cell surface antigen 2020373103   25 Mar 2026 is selected from the group consisting of EpCAM, E-cadherin, CD326, and any combination thereof. In some embodiments, the one cell type is a leukocyte and the cell surface antigen is CD45. In some embodiments, the one cell type is a mesenchymal cell and the cell surface antigen is selected from the group consisting of N-cadherin, OB-cadherin, alpha-5, beta-1 integrin, alpha-V, beta-6 integrin, Syndecan-1, and any combination thereof. In some embodiments, the method further comprises using an antibody that binds the cell surface antigen to separate the one cell type. In some embodiments, the fluid sample is disposed in a sample collector. In some embodiments, said sample collector is a pad, a tampon, a vaginal cup, a cervical cap, a menstrual disk, a cervical disk, a sponge, or an interlabial pad. In some embodiments, the sample collector comprises the buffer solution.

[0008] Disclosed herein, in some embodiments are methods for assessing menstrual health in a subject. In some embodiments, the method comprises: (a) receiving a menstrual fluid sample collected from said subject; (b) contacting said menstrual fluid sample with a buffer solution under conditions suitable to preserve a sample component selected from the group consisting of intact cells, nucleic acid, protein, and any combination thereof; and (c) determining a sample component level from the menstrual fluid sample. In some embodiments, the sample component comprises intact cells, and wherein step (b) maintains at least 90%, 95%, or substantially 100% of said one or more types of cells in a substantially intact state. In some embodiments, step (c) comprises determining an amount of one or more cell types in the menstrual fluid sample. In some embodiments, the one or more cell types are selected from the group consisting of leukocytes, erythrocytes, endothelial cells, epithelial cells, stromal cells, stem cells, and any combinations thereof. In some embodiments, the method further comprises comparing the amount of one or more cell types to a predetermined threshold. In some embodiments, an increase in the amount as compared to the predetermined threshold indicates that the subject has a menstrual cycle disorder. In some embodiments, the sample component comprises nucleic acid. In some embodiments, the method further comprises determining an amount of nucleic acid present in the menstrual fluid sample. In some embodiments, the method further comprises comparing the amount of nucleic acid to a predetermined threshold. In some embodiments, an increase in the amount as compared to the predetermined threshold indicates that the subject has a menstrual cycle disorder. In some embodiments, the predetermined threshold is an amount of nucleic acid present in a reference menstrual fluid sample. In some embodiments, the reference menstrual fluid sample is obtained from a healthy control subject. In some embodiments, the nucleic acid comprises RNA, and wherein the method further comprises measuring the level of 2020373103   25 Mar 2026 at least one RNA expression marker. In some embodiments, the nucleic acid comprises miRNA, and wherein the method further comprises measuring the level of at least one miRNA. In some embodiments, the method comprises comparing the level to a predetermined threshold. In some embodiments, the sample component comprises protein. In some embodiments, the method comprises determining the presence or level of at least one protein marker. In some embodiments, the method comprises comparing the level to a predetermined threshold. In some embodiments, the sample component comprises a microbial source present in the menstrual fluid sample. In some embodiments, the microbial source is a bacteria, a fungus, or a virus. In some embodiments, the method further comprises measuring a microbial source component, and wherein the component is selected from the group consisting of nucleic acid, protein, metabolite, cell, and any combination thereof. In some embodiments, the method further comprises measuring the bacterial diversity in the menstrual fluid sample. In some embodiments, said menstrual fluid sample is disposed in a sample collector. In some embodiments, said sample collector is a pad, a tampon, a vaginal cup, a cervical cap, a menstrual disk, a cervical disk, a sponge, or an interlabial pad. In some embodiments, the sample collector comprises the buffer solution. In some embodiments, said buffer solution has a pH greater than 7. In some embodiments, the method further comprises, prior to (c), storing said menstrual fluid sample in the presence of said buffer solution for at least 1 day. In some embodiments, the storage occurs at up to about 4 °C. In some embodiments, the storage occurs at up to about -20 °C. In some embodiments, the storage occurs at room temperature.

[0009] Disclosed herein in some embodiments, aremethods for detecting a menstrual cycle disorder in a subject. In some embodiments, the method comprises: (a) determining a flow rate of a menstrual fluid sample collected from said subject within a predefined time period; and (b) comparing said flow rate of said menstrual fluid sample to a predetermined threshold; wherein an increased flow rate relative to said predetermined threshold indicates that said subject has said menstrual cycle disorder. In some embodiments, said predetermined threshold is a flow rate of a menstrual fluid sample obtained from a reference subject within said predefined time period. In some embodiments, said reference subject is a healthy control subject. In some embodiments, said predefined time period is greater than 15 minutes. In some embodiments, said predefined time period is less than 2 hours. In some embodiments, the method further comprises measuring a volume of said menstrual fluid sample collected from said subject within said predefined time period. In some embodiments, the method further comprises comparing said measured volume to a predetermined threshold. In some embodiments, said predetermined 2020373103   25 Mar 2026 threshold is a volume of a menstrual fluid sample collected from a reference subject within said predefined time period. In some embodiments, said menstrual fluid sample is disposed in a sample collector. In some embodiments, said sample collector is a pad, a tampon, a vaginal cup, a cervical cap, a menstrual disk, a cervical disk, a sponge, or an interlabial pad.

[0010] Disclosed herein in some embodiments are methods for detecting a menstrual cycle disorder. In some embodiments, the method comprises: (a) determining an expression level of one or more markers in a fluid sample obtained from the vaginal cavity of a subject, wherein the one or more markers are selected from Table 1, Table 2 and / or Table 3; (b) applying a classifier algorithm to said expression level of one or more markers and a reference level of each of the one or more markers to calculate a metric that quantifies a difference between the expression level and the reference level for each of the one or more markers; and (c) determining a presence of a menstrual cycle disorder based on the metric. Disclosed herein, in some embodiments is a method of producing a desired preparation for assessment of menstrual health, comprising: (a) receiving a fluid sample collected from the vaginal cavity of a subject, the fluid sample comprising one or more types of cells; (b) contacting the fluid sample with a buffer solution under conditions suitable to maintain one or more cell types in a substantially intact state; (c) separating one cell type in the fluid sample from the remaining fluid sample; and (d) applying a classifier algorithm to said expression level of one or more markers in the one cell type to calculate a metric that quantifies the difference between said expression level and a reference level to assess a level of menstrual health, wherein the one or more markers are selected from Table 1 and / or Table 2.

[0011] Disclosed herein in some embodiments are methods for detecting a menstrual cycle disorder in a subject. In some embodiments, the method comprises: (a) determining a flow rate of a menstrual fluid sample collected from said subject within a predefined time period; and (b) applying a classifier algorithm to said flow rate of said menstrual fluid sample to calculate a metric that quantifies a difference between said metric and a predetermined threshold; wherein an increased flow rate relative to said predetermined threshold indicates that said subject has said menstrual cycle disorder.

[0012] Disclosed herein in some embodiments are methods comprising: (a) obtaining a fluid sample from a vaginal cavity of a subject, wherein the fluid sample is stabilized under conditions which preserve a component of the fluid sample;(b) determining an expression level of one or more markers in the fluid sample; (c) comparing said expression level to a reference level of said one or more markers to detect an increased or decreased expression level of the one 2020373103   25 Mar 2026 or more markers relative to the reference expression level, and (d) determining from the increased or decreased expression level whether the subject has a menstrual cycle disorder. In some embodiments, the component is preserved for at least 1 day. In some embodiments, the component is preserved at up to about 4 °C. In some embodiments, the component is preserved at up to about -20 °C. In some embodiments, the component is preserved at room temperature In some embodiments, the component is selected from the group consisting of a cell, a nucleic acid, a protein, a metabolite, and a microorganism. In some embodiments, the one or more markers are selected from the group consisting of the markers in Table 4. In some embodiments, the one or more markers are selected from the group consisting of the markers in Table 5. In some embodiments, the one or more markers are selected from the group consisting of the markers in Table 1. In some embodiments, the menstrual cycle disorder is HMB. In some embodiments, the menstrual cycle disorder is AUB. [0012A] A method of producing a desired preparation for assessment of menstrual health, comprising: (a) receiving a fluid sample collected from the vaginal cavity of a subject, the fluid sample comprising one or more components; (b) contacting the fluid sample, with a buffer solution under conditions to preserve one or more components of the fluid sample; (c) determining an expression level of a sample component from the one or more components of the fluid sample; (d) obtaining survey data from the first subject and from a group of subjects, wherein the survey data comprises one or more features selected from the group consisting of a menstrual cycle phenotype, a physical body characteristic, a disease or condition, a medical treatment, demographic information, lifestyle information, ancestry, sexuality, menstrual management, health care usage, health care access, and health record data relating to said group of subjects; and (e) stratifying the group of subjects into subgroups based on said one or more features; (f) assigning said first subject to one of said subgroups based on said one or more features; (g) comparing said expression level of step (c) to a reference expression level of the sample component to generate a comparison, wherein the reference expression level of the sample component is obtained from the subgroup to which said first subject was assigned; and (h) determining an assessment of menstrual health using the comparison obtained in step (g). [0012B] Throughout this specification the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated element, integer or step, or group of elements, integers or steps, but not the exclusion of any other element, integer or step, or group of elements, integers or steps. [0012C] Any discussion of documents, acts, materials, devices, articles or the like which has 2020373103   25 Mar 2026 been included in the present specification is not to be taken as an admission that any or all of these matters form part of the prior art base or were common general knowledge in the field relevant to the present disclosure as it existed before the priority date of each of the appended claims. INCORPORATION BY REFERENCE

[0013] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee. In some embodiments, the novel features of the invention are set forth with particularity in the appended claims. In some embodiments, a better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the invention are utilized, and the accompanying drawings of which:

[0015] FIG. 1 depicts a pattern of menstrual blood over the days of a cycle.

[0016] FIG. 2 depicts the different colors of menstrual blood.

[0017] FIG. 3 depicts the different sizes of tampons.

[0018] FIG. 4 depicts the different sizes of pads.

[0019] FIG. 5 depicts the different types of menstrual products used.

[0020] FIG. 6 depicts different hair loss patterns.

[0021] FIG. 7 depicts possible locations for acne on a subject.

[0022] FIG. 8 depicts acne severity.

[0023] FIG. 9 depicts different types of acne.

[0024] FIG. 10 depicts different patterns of hair on the upper lip.

[0025] FIG. 11 depicts different patterns of hair on the arms.

[0026] FIG. 12 depicts different patterns of hair on the jawline.

[0027] FIG. 13 depicts different patterns of hair on the thighs.

[0028] FIG. 14 depicts different patterns of hair on the chest.

[0029] FIG. 15 depicts different patterns of hair on the stomach. 2020373103   25 Mar 2026

[0030] FIG. 16 depicts different patterns of hair on the upper back.

[0031] FIG. 17 depicts different patterns of hair on the lower back.

[0032] FIG. 18 depicts different patterns of hair on the pubic area.

[0033] FIG. 19 depicts the location of the uterus.

[0034] FIG. 20 depicts a method to test pelvic floor strength.

[0035] FIG. 21 depicts locations where a subject may store fat.

[0036] FIG. 22 depicts a method to identify body size.

[0037] FIG. 23 depicts different types of caffeine.

[0038] FIG. 24 illustrates the diversity of bacterial species among patients with endometriosis, polycystic ovarian syndrome, and among healthy patients.

[0039] FIG. 25A-25C illustrate the different types of primary cell types, reproductive issues, and stem cells found in menstrual blood over time.

[0040] FIG. 26 displays the amount of alkaline hematin found in two patient samples of menstrual blood.

[0041] FIG. 27A-27C illustrate potential data groupings by stratified patient.

[0042] FIG. 28 illustrates factors used to annotate a patient.

[0043] FIG. 29 is a schematic of the comparison of a patient value over time as compared to the mean value of a other patients with similar annotations. DETAILED DESCRIPTION

[0044] Provided in this disclosure are methods for detecting a menstrual cycle disorder in a subject. In some embodiments, a subject is suspected of having or at risk for a menstrual cycle disorder. In some embodiments, a subject is a demographic, lifestyle, or genetic risk factor for a menstrual cycle disorder. In some embodiments, a subject exhibits expression of RNA, protein, metabolites or other cellular products that is linked as a risk factor for a menstrual cycle disorder. Menstrual Fluid Samples

[0045] In some embodiments of methods herein, fluid from the vaginal cavity of a subject is collected from the subject, such as by using a collection device. In some embodiments, the collection device is placed inside or near the vagina and left in place for a period of time to allow fluid such as menstrual fluid to collect in or on the device. In some embodiments, the fluid is then removed or extracted from the device for further analysis. In some embodiments, the fluid is menstrual fluid, such as menstrual blood collected during the subject’s menstrual 2020373103   25 Mar 2026 window. In some embodiments, fluid is collected from the vaginal cavity outside of the subject’s menstrual window.

[0046] In some embodiments, a menstrual cycle disorder is a disorder or abnormality associated with a menstrual cycle, which comprises heavy menstrual bleeding or abnormal bleeding, such as abnormal uterine bleeding. In some aspects, these methods comprise collecting a sample of fluid from the vaginal cavity of a subject (e.g., menstrual fluid) and analyzing an aspect of the fluid.

[0047] In some embodiments, aspects of fluid which are analyzed include, but are not limited to, flow rate, cell types, nucleic acid content, gene expression, protein biomarker expression, metabolites, and gene target expression. In some embodiments, these aspects are quantified, and in some cases, a change such as an increase or decrease in an aspect are correlated with a menstrual cycle disorder. In some embodiments, aspects of fluid which are analyzed include the biological markers (also referred to as “biomarkers”) described herein. In some embodiments, such a change are indicative of a menstrual cycle disorder.

[0048] In some cases, normal menstrual cycle occurs approximately every month and comprises the shedding of the lining of the uterus through the vagina. In some cases, normal menstrual flow lasts approximately 4 or 5 days, lasts up to 7 days, and occurs every 21 to 35 days.

[0049] In some embodiments, collection of fluid from the vaginal cavity, such as menstrual fluid, provides direct access to reproductive tissue. In some embodiments, this access allows for biopsy, biomarker detection, and assessment of health conditions. In some embodiments, knowledge gained from such information provides women with health information about reproductive conditions, which affects fertility or quality of life.

[0050] In some instances, menstrual cycle disorders are the most prevalent gynecologic health problems in the United States, and heavy menstrual bleeding (HMB) affects up to 30% of women at some time during their reproductive years. Many reproductive conditions present with changes to the menstrual cycle, for example, including light bleeding or heavy bleeding. In some embodiments, the menstrual cycle presents abnormal uterine bleeding (AUB). In some embodiments, AUB comprises bleeding or spotting between periods (e.g., off-cycle bleeding), bleeding or spotting after sex, heavy bleeding during menstruation, a menstrual cycle that is abnormally long (e.g., longer than 38 days), a menstrual cycle that is abnormally short (e.g., shorter than 24 days), irregular periods (e.g., periods in which the cycle length varies by at least 7, 8, or 9 days), or bleeding after menopause. In some instances, the societal and personal burden of AUB lies in its impact on quality of life, productivity, and health care use and costs. 2020373103   25 Mar 2026

[0051] In some embodiments, heavy menstrual bleeding is diagnosed using a pictorial blood loss assessment chart, such as the one depicted in FIG. 1.

[0052] In some embodiments, the alkaline haematin test provides a quantitative assessment for heavy menstrual bleeding. In some embodiments, this involves a collection of sanitary products in a menstrual cycle and soaking them in sodium hydroxide.

[0053] In some embodiments, one or more blood markers provides a measurement for heavy menstrual bleeding. In some embodiments, the blood marker is hematocrit, hemoglobin, zinc protoporphyrin, or any combination thereof. In some embodiments, the blood marker level or amount in a collected menstrual fluid sample is compared to the blood marker level or amount in a whole blood sample (e.g., venous blood) from the same subject.

[0054] In some instances, a menstrual cycle disorder is indicative of an underlying disease or disorder, which affect the health, quality of life, fertility, or lifespan of a woman. In some instances, menstrual cycle disorders and other disorders which present with, or cause, HMB or AUB include eating disorders; extreme weight loss; excessive exercise; polycystic ovary syndrome (PCOS); ovarian cysts; premature ovarian failure; breast cancer; ovarian cancer; infertility; diminished ovarian reserve; chronic or frequent urinary tract infections; ectopic pregnancy; heart disease; type 1 diabetes; type 2 diabetes; an autoimmune condition such as lupus, multiple sclerosis, or rheumatoid arthritis; pelvic inflammatory disease (PID); fibroids (e.g., uterine fibroids); adenomyosis; cervical cancer; endometrial cancer; uterine cancer; or infection of the cervix or endometrium. In some embodiments, HMB or AUB accompanies a disease affecting the kidney, liver, thyroid, or adrenal glands.

[0055] In some embodiments, fluid collected from the vaginal cavity is a biological matrix comprising a plurality of cell types. In some instances, the cell types include immune cells, ovarian cells, fallopian tube cells, endometrial cells, and cervical cells. In some embodiments, the fluid is menstrual fluid, including menstrual blood. In some instances, comparison of endometrium and other reproductive tissues from the fluid collected from the vaginal cavity, such as menstrual fluid, of healthy women and those with dysfunction advances understanding of key areas of endometrial physiology, including infertility, receptivity, endometriosis, and cancer. In some embodiments, a typical gene expression is linked to cellular dysfunction. In some instances, endometrial mucosa undergoes dynamic, hormone-dependent alterations throughout the life of females. In some instances, detailed information about menstrual cyclespecific gene expression changes promotes understanding of the gene regulatory networks which underlie changes in the menstrual cycle. In some embodiments, such changes lead to preparation 2020373103   25 Mar 2026 of uterus for embryo implantation. In further cases, such changes is analyzed to identify molecular differences between healthy and diseased endometrium.

[0056] In some instances, epigenetic factors such as methylation and the immune repertoire play a large role in initiating morphological and functional changes with the endometrium that promote or be essential for uterine receptivity. In some embodiments, these methylation changes is cycle dependent, and is associated with gene expression regulation. In some instances, the female reproductive tract comprise one or more immune cell populations, which is enriched for a particular cell type. In some embodiments, in some cases, an immune cell population becomes enriched in natural killer cells, which plays a role in endometrial receptivity or increase a risk for pregnancy loss.

[0057] New efforts have genomically characterized endometrium, allowing for single-cell resolution of tissue characterization for better disease genotyping. Studies have begun to shed light on the complex sample makeup of menstrual fluid, with a distinct immune repertoire. In some embodiments, eutopic endometrium or cells isolated from menstrual fluid has a genomic correlation with endometriosis.

[0058] Measurement of menstrual bleeding is one way to diagnose or monitor menstrual cycle disorders such as HMB or AUB. In some embodiments, heavy menstrual bleeding includes bleeding about 80 ml or more during a single menstrual period. However, almost half of all women reporting heavy menstrual bleeding has less than about 40 ml of menstrual bleeding per cycle. Thus, in some instances, accurate and reliable methods for quantifying menstrual bleeding, including heavy menstrual bleeding, is necessary to detect menstrual cycle disorders and to understand effects of HMB or AUB on health and discomfort of the patient.

[0059] In some embodiments, normal menstrual bleeding comprises a menstrual flow having a normal volume and flow rate. In some embodiments, normal menstrual bleeding presents with a typical expression of one or more genes, presence or absence of one or more cell types, a typical nucleic acid content, a typical flow rate, a typical protein biomarker expression, a metabolite, a hematocrit level, or a typical gene target biomarker expression.

[0060] In some embodiments, normal menstrual bleeding is in some cases be identified by a volume of menstrual fluid lost during a menstrual cycle or by a flow rate during a menstrual cycle. In some embodiments, normal menstrual bleeding comprises between 10 mL and 80 mL of menstrual blood during a single menstrual cycle. In some embodiments, normal menstrual bleeding comprises a maximum volume of at least 40 mL, at least 45 mL, at least 50 mL, at least 55 mL, at least 60 mL, at least 65 mL, at least 70 mL, at least 75 mL, at least 80 mL, at least 85 2020373103   25 Mar 2026 mL, at least 90 mL, at least 95 mL, or at least 100 mL of menstrual fluid lost during a menstrual cycle. In some embodiments, normal menstrual bleeding comprises a flow rate of up to 7 mL per day, up to 8 mL per day, up to 9 mL per day, up to 10 mL per day, up to 11 mL per day, up to 12 mL per day, up to 13 mL per day, up to 14 mL per day, up to 15 mL per day, up to 16 mL per day, up to 17 mL per day, up to 18 mL per day, up to 19 mL per day, or up to 20 mL per day. In some embodiments, normal menstrual bleeding comprises a flow rate of up to 0.05 mL per hour, up to 0.1 mL per hour, up to 0.2 mL per hour, up to 0.3 mL per hour, up to 0.4 mL per hour, up to 0.5 mL per hour, up to 0.6 mL per hour, up to 0.7 mL per hour, up to 0.8 mL per hour, up to 0.9 mL per hour, or up to 1.0 mL per hour.

[0061] In some cases, HMB is menstrual bleeding having a larger volume or higher flow rate than normal menstrual bleeding. In some cases, HMB present with an altered expression of one or more genes detectable in a menstrual fluid sample, presence or absence of one or more cell types detectable in a menstrual fluid sample, a high or low nucleic acid content detectable in a menstrual fluid sample, a high flow rate of menstrual fluid during at least a portion of a menstrual cycle, an altered protein or biomarker expression detectable in a menstrual fluid sample, an altered metabolite level in a menstrual fluid sample, an altered enzyme level in a menstrual fluid sample, an altered hematocrit level in a menstrual fluid level, or an altered gene target biomarker expression detectable in a menstrual fluid sample.

[0062] In some embodiments, HMB comprises at least 60 mL, at least 65 mL, at least 70 mL, at least 75 mL, at least 80 mL, at least 85 mL, at least 90 mL, at least 95 mL, at least 100 mL, at least 110 mL, at least 120 mL, or more of menstrual fluid lost during a single menstrual cycle. In some embodiments, HMB, such as HMB presenting as a high flow rate, comprises at least 30 mL, at least 40 mL, at least 50 mL, at least 60 mL, at least 70 mL, at least 80 mL, at least 90 mL, at least 100 mL, at least 110 mL, at least 120 mL, or more menstrual fluid lost during a single menstrual cycle. In some embodiments, HMB comprises a flow rate of at least 10 mL per day, at least 11 mL per day, at least 12 mL per day, at least 13 mL per day, at least 14 mL per day, at least 15 mL per day, at least 16 mL per day, at least 17 mL per day, at least 18 mL per day, at least 19 mL per day, or at least 20 mL per day of menstrual fluid lost during at least a portion of a single menstrual cycle. In some embodiments, HMB comprises a flow rate of at least 0.1 mL per hour, at least 0.2 mL per hour, at least 0.3 mL per hour, at least 0.4 mL per hour, at least 0.5 mL per hour, at least 0.6 mL per hour, at least 0.7 mL per hour, at least 0.8 mL per hour, at least 0.9 mL per hour, at least 1.0 mL per hour, at least 1.1 mL per hour, at least 1.2 mL per hour, at least 1.3 mL per hour, at least 1.4 mL per hour, or at least 1.5 mL per hour of menstrual fluid 2020373103   25 Mar 2026 lost during a single menstrual cycle. In some embodiments, HMB, such as HMB presenting as a high flow rate, comprises a volume of menstrual fluid lost that is consistent with normal menstrual bleeding. In such cases, a volume of menstrual fluid lost is at least about 30 mL, 40 mL, 50 mL, 60 mL, 70 mL, 80 mL, 90 mL, 100 mL, 110 mL, 120 mL, or more menstrual fluid during a single menstrual cycle. In some embodiments, HMB lasts for up to 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9 ,10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 30, 29, or 30 days. In some embodiments, HMB lasts for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 30, 29, 30, or more days. HMB lasting longer than 7 days is menorrhagia.

[0063] AUB is menstrual bleeding having a volume or flow rate that is different than that of normal menstrual bleeding. In some embodiments, AUB is HMB. In some embodiments, AUB presents with an altered expression of one or more genes detectable in a menstrual fluid sample, presence or absence of one or more cell types detectable in a menstrual fluid sample, a high or low nucleic acid content detectable in a menstrual fluid sample, a high flow rate of menstrual fluid during at least a portion of a menstrual cycle, an altered protein or biomarker expression detectable in a menstrual fluid sample, an altered metabolite level in a menstrual fluid sample, an altered hematocrit level in a menstrual fluid level, or an altered gene target biomarker expression detectable in a menstrual fluid sample.

[0064] In some embodiments, AUB comprises at least 20 mL, at least 30 mL, at least 40 mL, at least 50 mL, at least 60 mL, at least 70 mL, at least 80 mL, at least 90 mL, at least 100 mL, at least 110 mL, at least 120 mL, or more menstrual fluid lost during a single menstrual cycle. In some embodiments, AUB comprises a flow rate of at least 7 mL per day, at least 8 mL per day, at least 9 mL per day, 10 mL per day, at least 11 mL per day, at least 12 mL per day, at least 13 mL per day, at least 14 mL per day, at least 15 mL per day, at least 16 mL per day, at least 17 mL per day, at least 18 mL per day, at least 19 mL per day, or at least 20 mL per day of menstrual fluid lost during at least a portion of a single menstrual cycle. In some embodiments, AUB comprises a flow rate of at least 0.1 mL per hour, at least 0.2 mL per hour, at least 0.3 mL per hour, at least 0.4 mL per hour, at least 0.5 mL per hour, at least 0.6 mL per hour, at least 0.7 mL per hour, at least 0.8 mL per hour, at least 0.9 mL per hour, at least 1.0 mL per hour, at least 1.1 mL per hour, at least 1.2 mL per hour, at least 1.3 mL per hour, at least 1.4 mL per hour, or at least 1.5 mL per hour of menstrual fluid lost during at least a portion of a single menstrual cycle. In some embodiments, AUB, such as AUB presenting as a high flow rate, comprises a volume of menstrual fluid lost that is consistent with normal menstrual bleeding. In such cases, a 2020373103   25 Mar 2026 volume of menstrual fluid lost is at least about 30 mL, 40 mL, 50 mL, 60 mL, 70 mL, 80 mL, 90 mL, 100 mL, 110 mL, 120 mL, or more menstrual fluid during a single menstrual cycle. In some embodiments, AUB lasts for up to 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 days. In some embodiments, AUB lasts for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or more days. In some embodiments, AUB is HMB.

[0065] In some embodiments, HMB presents as an alteration in the distribution, ratio or amounts of cell types in menstrual fluid. For example, HMB may present as an increase in red blood cells in the menstrual fluid of an HMB subject, or an alteration in the ratios between endothelial, epithelial and hematopoietic cells or any combination thereof.

[0066] Described herein are methods for detecting a menstrual cycle disorder in a subject. Briefly, a fluid sample is obtained from the vaginal cavity of a subject for analysis, and a property of the fluid is determined. In some embodiments, properties of the fluid sample comprises (i) gene expression detectable in a fluid sample, (ii) presence or absence of a cell type detectable in a fluid sample, (iii) amount of nucleic acid detectable in a fluid sample, (iv) flow rate of menstrual fluid during a portion or all of a menstrual cycle, (v) one or more protein biomarkers detectable in a fluid sample, (vi) one or more metabolite detectable in a fluid sample, (vii) one or more enzyme detectable in a fluid sample, and / or (viii) one or more gene target biomarkers detectable in a fluid sample. Such properties of a fluid sample is detected or measured in a fluid sample from the vaginal cavity during the menstrual window and / or outside the menstrual window. In some embodiments, samples is from a subject experiencing menstrual bleeding, such as normal menstrual bleeding, HMB, or AUB.

[0067] In some embodiments, a fluid sample, such as a menstrual fluid sample or a sample of another fluid, is collected from a subject using a sample collector which collects fluid from the vaginal cavity. In some embodiments, a sample collector is placed in the vagina or outside the vagina for sample collection. In some embodiments, a sample collector collects a sample by pooling, holding, catching, directing, or absorbing the sample. In some embodiments, a sample collector is absorbent, semi-absorbent, or non-absorbent. In some embodiments, a sample collector is soluble in a buffer. In some embodiments, a sample collector is broken down, for example by exposing the sample collector to an acidic environment, a basic environment, or an enzyme. In some embodiments, sample collectors comprise a pad, a tampon, a vaginal cup, a cervical cap, a menstrual disk, a cervical disk, a sponge, or an interlabial pad. In some embodiments, more than one type of sample collector are used. 2020373103   25 Mar 2026

[0068] In some embodiments, a sample collector is left in place for a pre-determined amount of time to collect a fluid sample. In some embodiments, at least 5 minutes, 10 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 1.5 hours, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, or 8 hours elapses. In some embodiments, at most 5 minutes, 10 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 1.5 hours, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, or 8 hours elapses while the sample collection device is left in place. In some embodiments, about 5 minutes, 10 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 1.5 hours, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, or 8 hours elapses while the sample collector is left in place

[0069] In some embodiments, a sample is collected during the menstrual window (the period) of a subject. In some embodiments, a sample collector is disposable. In some embodiments, a disposable sample collector is discarded or broken down after use. In some embodiments, a disposable sample collector is dissolvable, biodegradable, recyclable, or compostable. Typically, one disposable sample collector is used to collect one sample from one subject. In some embodiments, a sample collector is reusable. In some embodiments, a reusable sample collector is washable, sterilizable, or autoclavable. In some embodiments, a reusable sample collector is resistant to degradation, tearing, pore formation, or dissolution. In some embodiments, a reusable sample collector comprises anti-microbial, antibacterial, antiviral, or antifungal properties. In some embodiments, a reusable sample collector is used one or more times to collect one or more samples. In some embodiments, a reusable sample collector is used about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, or more times to collect one or more fluid samples. In some embodiments, a reusable sample collector is used to repeatedly collect fluid samples from one subject. In some embodiments, a reusable sample collector is used to collect samples from a plurality of subjects.

[0070] In some embodiments, one or more sample is collected during one or more periods (menstrual windows) of a subject. In some embodiments, 1 sample is collected during 1 period cycle, 2 samples is collected during 1 period cycle, 3 samples is collected during 1 period cycle, 4 samples is collected during 1 period cycle, more than 4 samples is collected during 1 period cycle, 2 samples is collected during 2 period cycles, 3 samples is collected during 2 period cycles, 4 samples is collected during 2 period cycles, 5 samples is collected during 2 period cycles, 6 samples is collected during 2 period cycles, 7 samples is collected during 2 period cycles, 8 samples is collected during 2 period cycles, more than 8 samples is collected during 2 period cycles, 3 samples is collected during 3 period cycles, 4 samples is collected during 3 period cycles, 5 samples is collected during 3 period cycles, 6 samples is collected during 3 2020373103   25 Mar 2026 period cycles, 7 samples is collected during 3 period cycles, 8 samples is collected during 3 period cycles, 9 samples is collected during 3 period cycles, 10 samples is collected during 3 period cycles, 11 samples is collected during 3 period cycles, 12 samples is collected during 3 period cycles, more than 12 samples is collected during 3 period cycles, 4 samples is collected during 4 period cycles, 5 samples is collected during 4 period cycles, 6 samples is collected during 4 period cycles, 7 samples is collected during 4 period cycles, 8 samples is collected during 4 period cycles, 9 samples is collected during 4 period cycles, 10 samples is collected during 4 period cycles, 11 samples is collected during 4 period samples, 12 samples is collected during 4 period cycles, 13 samples is collected during 4 period cycles, 14 samples is collected during 4 period cycles, 15 samples is collected during 4 period cycles, 16 samples is collected during 4 period cycles, or more than 16 samples is collected during 4 period cycles. In some embodiments, a plurality of samples is collected during more than 4 period cycles.

[0071] In some embodiments, a plurality of samples is collected during a single period cycle, and in some cases, in a single day. For example, samples are collected for different durations of collection on a single day or on multiple days, such as collections for 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5 hours or 6 hours.

[0072] In some embodiments, samples are collected outside the menstrual window, e.g., between the time of the subject’s periods. In such cases, a non-menstrual fluid is collected using the sample collector. In some embodiments, non-menstrual fluid which is collected include vaginal secretions, cervical mucus, cervicovaginal fluid, spotting blood (i.e., from between periods), amniotic fluid, a mucus plug, or other vaginal discharge. In some embodiments, non-menstrual fluid is collected and analyzed using a protocol which is used to collect and analyze menstrual fluid.

[0073] In some embodiments, a sample is collected after a menstrual window has closed, e.g., after a period has ended. In some embodiments, a sample is collected on the same day a menstrual window closed. In some embodiments, a sample is collected about 1 day, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, about 7 days, about 8 days, about 9 days, about 10 days, about 11 days, about 12 days, about 13 days, about 14 days, about 15 days, about 16 days, about 17 days, about 18 days, about 19 days, about 20 days, about 21 days, about 22 days, about 23 days, about 24 days, about 25 days, about 26 days, about 27 days, about 28 days, about 29 days, or about 30 days after a menstrual window has closed. In some embodiments, a sample is collected at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 8 days, at least 9 days, at least 10 days, at 2020373103   25 Mar 2026 least 11 days, at least 12 days, at least 13 days, at least 14 days, at least 15 days, at least 16 days, at least 17 days, at least 18 days, at least 19 days, at least 20 days, at least 21 days, at least 22 days, at least 23 days, at least 24 days, at least 25 days, at least 26 days, at least 27 days, at least 28 days, at least 29 days, or at least 30 days after a menstrual window has closed. In some embodiments, sample is collected not more than 1 day, not more than 2 days, not more than 3 days, not more than 4 days, not more than 5 days, not more than 6 days, not more than 7 days, not more than 8 days, not more than 9 days, not more than 10 days, not more than 11 days, not more than 12 days, not more than 13 days, not more than 14 days, not more than 15 days, not more than 16 days, not more than 17 days, not more than 18 days, not more than 19 days, not more than 20 days, not more than 21 days, not more than 22 days, not more than 23 days, not more than 24 days, not more than 25 days, not more than 26 days, not more than 27 days, not more than 28 days, not more than 29 days, or not more than 30 days after a menstrual window has closed. In some embodiments, a sample is collected between 1 day and 30 days, between 1 day and 25 days, between 1 day and 20 days, between 1 day and 15 days, between 1 day and 10 days, between 1 day and 5 days, between 5 days and 30 days, between 5 days and 25 days, between 5 days and 20 days, between 5 days and 15 days, between 5 days and 10 days, between 10 days and 30 days, between 10 days and 25 days, between 10 days and 20 days, between 10 days and 15 days, between 15 days and 30 days, between 15 days and 25 days, between 15 days and 20 days, between 20 days and 30 days, between 20 days and 25 days, or between 25 days and 30 days after a menstrual window has closed.

[0074] In some embodiments, non-menstrual fluid collected between two menstrual windows is collected during various points during the reproductive cycle. In some embodiments, non-menstrual fluid is collected during a pre-ovulation phase, during ovulation, or during a postovulation phase. In some embodiments, non-menstrual fluid is collected during a proliferative phase, or during a luteal or secretory phase. In some embodiments, a phase of the reproductive cycle is an abnormal phase. In some embodiments, menstrual fluid and non-menstrual fluid is collected from the same subject.

[0075] In some embodiments, a sample is collected between two menstrual windows. In some embodiments, a sample is collected about halfway between two menstrual windows, before the halfway point between two menstrual windows, or after the halfway point between two menstrual windows.

[0076] In some embodiments, multiple samples are collected between two menstrual windows. In some embodiments, 2, 3, 4, 5, 6, 7, or 8 samples is collected between two menstrual windows. 2020373103   25 Mar 2026 In some such cases, the multiple samples is collected from different times between the two menstrual windows.

[0077] In some embodiments, a sample is collected between two menstrual windows, while a second sample is collected between a second two menstrual windows. In further cases, a third sample is collected between a third two menstrual windows. In a general case, an nth sample is collected between n two menstrual windows, where n is a positive integer which is equal to 1 or more.

[0078] In some embodiments, fluid samples are collected from a subject both during a menstrual window and between a menstrual window. In some embodiments, a fluid sample is collected from a subject during a menstrual window, and a second fluid sample is collected from the same subject between two menstrual windows. In some embodiments, a fluid sample is collected from a subject during a menstrual window and a second fluid sample is collected from the same subject after the end of that menstrual window, and before the next menstrual window. In some embodiments, a fluid sample is collected from a subject before the start of a menstrual window, and a second fluid sample is collected from the same subject during that menstrual window.

[0079] In some embodiments, a volume of fluid, such as menstrual fluid or other fluid collected from a vaginal cavity, is determined using the sample collector. In some embodiments, a volume of menstrual fluid in a sample collector is determined for example by reading graduations on the sample collector. In some embodiments, graduations are at least 0.01 mL, 0.02 mL, 0.03 mL, 0.04 mL, 0.05 mL, 0.06 mL, 0.07 mL, 0.08 mL, 0.09 mL, 0.1 mL, 0.2 mL, 0.3 mL, 0.4 mL, 0.5 mL, 0.6 mL, 0.7 mL, 0.8 mL, 0.9 mL, or 1.0 mL. In some embodiments, a volume of menstrual fluid in a sample collector is determined by measuring the mass of fluid inside the sample collector. In some embodiments, a volume of menstrual fluid in a sample collector is determined for example by compressing or otherwise expelling the fluid from the sample collector and measuring the volume (or mass) of the expelled fluid.

[0080] In some embodiments, collected fluid such as menstrual fluid is extracted from the sample collector. In some embodiments, extraction occur by pouring, pipetting, or suctioning of the fluid, which is appropriate, for example, when the sample collector comprises a menstrual cup or other non-absorbent reservoir. In some embodiments, extraction occur by dissolving or otherwise breaking down and removing the sample collector from the sample, which is appropriate, for example, when the sample collector comprises a sponge, a tampon, a pad, or another absorbent material. 2020373103   25 Mar 2026

[0081] Provided herein are also methods for collecting and preserving a sample such as a sample of fluid collected form a vaginal cavity (e.g., menstrual fluid). In some embodiments, such methods is used to accurately and / or reliably indicate a menstrual disorder, such as HMB or AUB, or another such disorder.

[0082] In some embodiments, such methods comprise collection of a sample of fluid from the vaginal cavity of a subject. In some embodiments, collection is for example be performed using a collection device such as a sponge, a tampon, a pad, or another absorbent material. In some embodiments, a further description of collection devices and methods is provided herein.

[0083] In some embodiments, the fluid is stabilized. In some embodiments, stabilization comprises adding the fluid to a buffer, such as a preservation buffer. In some embodiments, stabilization also comprises storage at a specific temperature (e.g., 4 °C, -20 °C, room temperature, or another acceptable temperature, including those described herein). In some embodiments, stabilization comprises moving the sample to a new vessel and placing a lid or covering on the vessel. In some embodiments, the vessel is vacuum sealed, or the sample is stored under argon gas or nitrogen gas.

[0084] In some embodiments, preservation and stabilization by such methods preserves a component of the sample, such that the component remains substantially unchanged, or that the component does not deteriorate significantly while the sample is stored. In some embodiments, components include a cell, a nucleic acid, a gene target biomarker, a protein biomarker, another protein, a microorganism (e.g., a pathogen or member of a microbiome), a small molecule, a metabolite, or another component.

[0085] In some embodiments, a component or property of a component is measured, either quantitatively or qualitatively. In some embodiments, an expression level of a marker is determined, an amount of a metabolite is measured, a small molecule is quantified, a microorganism is identified, a protein is measured, or a nucleic acid is quantified.

[0086] In some embodiments, the measured property or component is compared to a reference level. In some embodiments, a reference level is a level of the component or property in a sample from a vaginal cavity of a subject not experiencing HMB or AUB, or of a subject who is healthy. In some embodiments, an increased or decreased expression level of the one or more markers relative to the reference expression level. In some embodiments, such an increase or decrease indicates HMB or AUB. In some embodiments, the method comprises determining from the increased or decreased expression level whether the subject has a menstrual cycle disorder. 2020373103   25 Mar 2026 Gene Expression

[0087] In some embodiments of methods herein, fluid from the vaginal cavity, such as menstrual fluid comprises a gene expression profile. In some embodiments, during HMB or AUB, this gene expression profile is altered. In some embodiments, alterations to the gene expression profile comprises an increase and / or a decrease in the expression of one or more genes in females having HMB or AUB compared with the menstrual fluid of females having normal menstrual bleeding.

[0088] In some embodiments, expression of certain genes in endothelial cells from an endometrium of a female diagnosed with HMB display a reduction in expression of UEA-1 and CD31 and overexpression of F8RA and CD34 when compared with endothelial cells from an endometrium of a female having normal menstrual bleeding. In some embodiments, distinct patterns of expression of osteopontin, laminin, fibronectin, and collagen IV is observed in a female with HMB. In some embodiments, an increase in endothelial cell proliferation occur in in the endometrium of a female with HMB.

[0089] In some methods, expression of one or more genes in a sample of fluid collected from a vaginal cavity menstrual fluid is detected or measured. In some embodiments, genes measured is genes which are normally expressed in fluid such as menstrual fluid or genes which are not normally expressed in fluid such as menstrual fluid. In some embodiments, expressed genes which are measured is genes which is expressed or absent during HMB or AUB. Genes has a higher or lower expression during HMB than during normal menstrual bleeding. In some embodiments, such genes include UEA-a, CD31, F8RA, CD34, osteopontin, laminin, fibronectin, collagen IV, or expression products thereof.

[0090] In some embodiments, expression or accumulation of protein and / or RNAs is upregulated during HMB or AUB such as, for example, one or more of HLA-Aa (Major histocompatibility complex class I), IL-6ST (Interleukin 6 signal transducer), APCDD1L (Adenomatosis polyposis coli downregulated 1-like ), TBX3 (T-box 3 ), UBN1a (Ubinuclein), DSPa (Desmoplakin) , SDCBP2 (Syndecan binding protein (syntenin)) , EZH2 (Enhancer of zeste 2 polycomb repressive complex 2 subunit) , UHMK1 (U2AF homology motif (UHM) kinase), NR2C2 (Nuclear receptor subfamily 2, group C, member 2), MIR1282a (microRNA), TMED6a (Transmembrane p24 trafficking protein), VAV3 (Vav 3 guanine nucleotide exchange factor), CDC42BPA (CDC42 binding protein kinase alpha (DMPK-like)) , C17orf75 (Chromosome 17 open reading frame), MB21D1 (Mab-21 domain containing 1), PTPRC (Protein tyrosine phosphatase, receptor type C), WISP1 (WNT1 inducible signaling pathway 2020373103   25 Mar 2026 protein 1), CDC27 (Cell division cycle 27), and FSD1L (Fibronectin type III and SPRY domain containing 1-like).

[0091] In some embodiments, expression or accumulation of protein and / or RNAs is downregulated during HMB or AUB such as, for example, one or more of BPIFB1 ((C20orf114)a BPI fold containing family B, member 1), SCGB3A1a (Secretoglobin, family 3A, member 1) TFF3a (Trefoil factor 3 (intestinal)), SCGB1D2a (Secretoglobin, family 1D, member 2), SCGB2A2a (Secretoglobin, family 2A, member 2), PRODH (Proline dehydrogenase (oxidase) 1), MFF (Mitochondrial fission factor), TSPAN8 (Tetraspanin), CXCL6a (Chemokine (C-X-C motif) ligand 6), SPDYE2 (Speedy / RINGO cell cycle regulator family member E2), FCGBP (Fc fragment of IgG binding protein), IRX6 (Iroquois homeobox 6), ADAM10 (ADAM metallopeptidase domain 10), MUC5ACa (Mucin 5AC, oligomeric mucus / gel-forming), TRHDE-AS1 (TRHDE antisense RNA (LOC283392)), CYP26A1 (Cytochrome P450, family 26, subfamily A, polypeptide 1), SAA2a (Serum amyloid A2), MMEL1 (Membrane metalloendopeptidase-like 1), GRIP2 (Glutamate receptor interacting protein 2), and SAA1a (Serum amyloid A1).

[0092] In some embodiments, genes expressed in a sample of fluid from the vaginal cavity, such as menstrual fluid, is measured using an acceptable method. In some embodiments, methods for measuring gene expression include polymerase chain reaction (PCR), quantitative real-time (q-RT-PCR), isothermal PCR, restriction enzyme analysis, RNA sequencing (e.g., sequencing mRNA), northern blotting, serial analysis of gene expression (SAGE), in situ hybridization (ISH), fluorescence ISH (FISH), microarray analysis.

[0093] In some embodiments, PCR-based systems use consensus or degenerate primer sequences to allow for amplification and identification of expressed RNA sequences.

[0094] In some embodiments, the presence or absence of an expressed gene is measured. Presence of an expressed gene is determined, for example, by detecting at least a trace amount of a gene in a sample. In some embodiments, an expressed gene is present if it is detectable after about 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, or 40 cycles of PCR. In some embodiments, an expressed gene is present if it is detectable by sequencing. In some embodiments, an expressed gene is present if a detectable signal is produced in response to the expressed gene in an imaging experiment such as ISH or FISH. In some embodiments, at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 100, 150, 200, 300, 400, 500, 1000, or more copies of an expressed gene is detectable. In some embodiments, absence of an expressed gene is determined, for example, as failure to detect a gene after about 30, 31, 32, 33, 34, 45, 56, 37, 38, 39, or 40 cycles. In some 2020373103   25 Mar 2026 embodiments, absence of an expressed gene is determined if it is not detected by sequencing in at least 1, 2, 3, 4, 5, or more samples. In some embodiments, absence of an expressed gene is determined if no detectable signal is produced in response to the expressed gene in an imaging experiment, such as ISH or FISH. In some embodiments, an expressed gene is absent if there are no copies in a sample of menstrual fluid. In some embodiments, an expressed gene is absent if the gene is expressed at a level below a threshold, such as a threshold of detection.

[0095] In some embodiments, gene expression in a sample is increased or decreased compared with a predetermined threshold. In some embodiments, a predetermined threshold of gene expression is determined from a reference sample, such as a reference sample collected from a subject experiencing normal menstrual bleeding, a subject experiencing HMB, or a subject experiencing AUB. In some embodiments, a threshold gene expression is an upper threshold, such as the maximum gene expression measured during normal menstrual bleeding. In some embodiments, a threshold gene expression is a lower threshold, such as the minimum gene expression measured during normal menstrual bleeding. In some embodiments, a threshold of gene expression is determined by measuring gene expression of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing normal menstrual bleeding. In some embodiments, a threshold of gene expression is determined by measuring the gene expression of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing HMB. In some embodiments, a threshold of gene expression is determined by measuring the gene expression of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing AUB. In some embodiments, a threshold of a gene expression is determined in a sample of fluid collected from a vaginal cavity other than menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects. In some embodiments, a threshold of gene expression is determined by comparing the gene expression of a sample of menstrual fluid of subjects experiencing normal menstrual bleeding and HMB, or subjects experiencing normal menstrual bleeding and AUB. In some embodiments, a threshold of gene expression is an average gene expression, a mean gene expression, a mode gene expression, a maximum gene expression, a minimum gene expression, an average gene expression plus 1, 2, or 3 standard deviations of gene expression, or an average gene expression minus 1, 2, or 3 standard deviations of gene expression.

[0096] In some embodiments, gene expression is measured as an absolute expression or a relative expression. In some embodiments, absolute expression is measured as an amount of gene present in a tested sample, and is expressed for example as a number of copies, as a mass 2020373103   25 Mar 2026 (e.g., mg), as a number of copies per volume, or as a mass per volume in a menstrual fluid sample. In some embodiments, relative expression is measured as an amount of a gene normalized to another value. In some embodiments, relative expression is normalized for example to a total amount of gene expression in a menstrual fluid sample, an amount of menstrual fluid in a menstrual fluid sample, an amount of cells in a menstrual fluid sample, an amount of a cell type in a menstrual fluid sample, to an amount of a protein biomarker in a menstrual fluid sample, to an amount of a gene target biomarker in a menstrual fluid sample, or to an expression of a same gene in a menstrual fluid sample from a subject experiencing normal menstrual bleeding.

[0097] In some embodiments, gene expression is compared to the expression level of the gene in a sample of fluid collected from a vaginal cavity such as menstrual fluid collected from a reference subject. In some embodiments, expression of a gene is increased or decreased in a sample of a subject experiencing HMB or AUB compared with a sample of a subject experiencing normal menstrual bleeding.

[0098] In some embodiments, absolute and / or relative expression levels of a gene measured using the methods herein is increased or decreased relative to normal levels of that gene. In some embodiments, this normal gene expression level is a reference expression level. In some embodiments, a reference expression level is the expression of a same gene in the fluid collected from the vaginal cavity of a control subject, such as menstrual fluid of a woman experiencing normal menstrual bleeding. In some embodiments, a reference expression level is the expression of a housekeeping gene, such as gapdh or P-actin. In some embodiments, using a reference expression level, one determines whether the expression of a given gene is increased or decreased in a sample.

[0099] In some embodiments, gene expression is measured on any day a subject is experiencing menstrual bleeding. In some embodiments, gene expression is measured more than once during menstrual bleeding. In some embodiments, gene expression is measured on day 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 of menstrual bleeding. Sometimes, gene expression is measured after 30 days of menstrual bleeding. In some embodiments, gene expression is measured on a day a subject is not experiencing menstrual bleeding, such as between two menstrual windows.

[0100] In some samples, such as samples from a woman experiencing HMB or AUB, one or more genes is increased or decreased compared with a reference sample. In some embodiments, an increase in a gene expression is indicative of HMB, AUB, a menstrual cycle disorder, or 2020373103   25 Mar 2026 another disorder. In some embodiments, a decrease in a gene expression is indicative of HMB, AUB, a menstrual cycle disorder, or another disorder.

[0101] In some embodiments, expression of one or more genes in a sample of a subject experiencing HMB or AUB compared with a sample of a subject experiencing normal menstrual bleeding is correlated or anti-correlated with the presence or severity of HMB or AUB. In some embodiments, an increase in gene expression is correlated with HMB or an increase in gene expression is correlated with AUB. In some embodiments, a decrease in gene expression is correlated with HMB or a decrease in gene expression is correlated with AUB. In some embodiments, an increase of expression of a first gene and a decrease expression of a second gene is correlated with HMB or AUB.

[0102] In some embodiments, expression of certain genes present at a given ratio in a normal sample is present in an increased or decreased ratio in a sample of a subject experiencing HMB or AUB. In some embodiments, the ratio of two genes is increased in a sample of a subject experiencing HMB or AUB compared with the ratio of the same two genes in a sample of a subject experiencing normal menstrual bleeding. In some embodiments, the ratio of two genes is decreased in a sample of a subject experiencing HMB or AUB compared with the ratio of the same two genes in a sample of a subject experiencing normal menstrual bleeding. Cell types

[0103] In some embodiments of methods herein, fluid taken from the vaginal cavity is a complex, heterogeneous mixture of one or more cell populations and comprises one or more cell types. During HMB or AUB, the cell types found in fluid, such as menstrual blood, is altered. In some embodiments, a fluid sample from the vaginal cavity of a subject experiencing HMB or AUB comprises a different composition of cell types, more of at least one cell type, or less of at least one cell type than a subject experiencing normal menstrual bleeding.

[0104] In some embodiments, a cell type in a fluid sample from the vaginal cavity of a subject, such as menstrual fluid, comprises a leukocyte, an erythrocyte, an endothelial cell, an epithelial cell, a stromal cell, a stem cell, a mesenchymal cell, or a combination thereof. In some embodiments, a cell type is present in a sample from a subject experiencing HMB or AUB that is absent in a sample from a subject experiencing normal menstrual bleeding. In some embodiments, a cell type is absent in a sample from a subject experiencing HMB or AUB that is present in a sample from a subject experiencing normal menstrual bleeding. In some embodiments, a cell type is present in a higher quantity in a sample from a subject experiencing HMB or AUB than in a sample from a subject experiencing normal menstrual bleeding. In some 2020373103   25 Mar 2026 embodiments, a cell type is present in a lower quantity in a sample from a subject experiencing HMB or AUB than in a sample from a subject experiencing normal menstrual bleeding.

[0105] In some embodiments, a cell type is determined based on the presence, absence, or expression of a cell surface antigen. In some embodiments, a cell type is determined based on a combination or relative expression of two or more cell surface antigens. In some embodiments, a cell is an endothelial cell. In some embodiments, cell surface antigens includes, CD31 / PECAM-1, CD34, CD36 / SR-B3, CD39, CD44, CD47, CD54 / ICAM-1, CD61, CD62E, CD62P, CD80, CD86, CD93, CD102, CD105, CD106, CD112, CD117, ESAM, ENDOMUCIN, CXCL16, CD121a, CD141, CD142, CD143, CD144, CD146, CD147, CD151, CD160, CD201, CD213a, CD248, CD309, ADAMS 8-17, 33, ADAMTS-13, ADAMTS-18, VWF, TEM8, NOTCH, KLF4, or a combination thereof. In some embodiments, a cell is an epithelial cell. In some embodiments, cell surface antigens include, for example, EpCAM, E-cadherin, CD326, or a combination thereof. In some embodiments, a cell is a leukocyte. In some embodiments, cell surface antigens include CD45. In some embodiments, a cell is a mesenchymal cell. In some embodiments, cell surface antigens include, for example, N-cadherin, OB-cadherin, alpha-5, beta-1 integrin, alpha-V, beta-6 integrin, Syndecan-1, or a combination thereof. In some embodiments, a cell type comprises at least 1, at least 2, at least 3, at least 4, at least 5, at least 10, at least 15, or at least 20 cell surface antigens such as those disclosed herein. In some embodiments, a cell type comprises no more than 1, no more than 2, no more than 3, no more than 4, no more than 5, no more than 10, no more than 15, or no more than 20 cell surface antigens such as those disclosed herein.

[0106] In some embodiments, a cell type present in a menstrual sample is identified from menstrual fluid collected on any day a subject is experiencing menstrual bleeding. In some embodiments, cell types is identified in samples taken at more than one time point during the menstrual cycle. Cell types is identified in samples taken on day 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 of menstrual bleeding. Sometimes, a cell type is identified in a menstrual fluid sample collected after 30 days of menstrual bleeding. In some embodiments, a cell type is identified when a subject is not experiencing menstrual bleeding, such as between two menstrual windows.

[0107] In some embodiments, a cell type of a cell in a sample is determined by an acceptable method, which includes, but is not limited to, flow cytometry, immunohistochemistry, immunocytochemistry, gene expression analysis, protein expression analysis, or metabolome 2020373103   25 Mar 2026 analysis of the cell. In some embodiments, at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, or more cell types is determined.

[0108] In some embodiments, normalization and cellular characterization analysis is used for identification of cell types. In some embodiments, such analysis requires granular genomic analysis or bioinformatic deconvolution.

[0109] In some embodiments, an amount of a cell type in a sample is determined by measuring the amount of that cell type using an acceptable method, which includes flow cytometry, immunohistochemistry, immunocytochemistry, gene expression analysis, protein expression analysis, or metabolome analysis of a plurality of cells. In some embodiments, an amount of a cell type is determined using whole sample. In some embodiments, an amount of a cell type is determined by measuring an amount of a type of cells in a subset of cells of the sample, such as a subset of cells that is removed from the whole sample.

[0110] In some embodiments, a presence of a cell type is determined. In some embodiments, a presence of a cell type is expressed as yes or no, wherein yes indicates that at least one of a cell type is identified in a menstrual fluid sample, and no indicates that none of a cell type is identified in a menstrual fluid sample. In some embodiments, a presence of a cell type is expressed semi-quantitatively (e.g. no expression, low expression, moderate expression, or high expression).

[0111] In some embodiments, an amount of a cell type is a total amount or a normalized amount of that cell type. In some embodiments, an amount is normalized for example to the total amount of cells in a sample, to a total amount of a subset of cells in the sample, to a biomarker, to a volume, to an amount of time. In some embodiments, an amount of a cell type is a relative amount of that cell type compared with another type of cell in the same sample. In some embodiments, an amount of a cell type is a relative amount of that cell type compared with the amount of the same cell type in a second sample. In some embodiments, the second sample is for example a sample of a woman experiencing a normal menstrual period.

[0112] In some embodiments, an amount of a cell type is compared with a predetermined threshold. In some embodiments, an amount of a cell type is increased or decreased by a predetermined threshold. In some embodiments, a predetermined threshold of an amount of a cell type is determined from a reference sample, such as a fluid collected from the vaginal cavity of a control subject, such as menstrual fluid collected from a subject experiencing normal menstrual bleeding, a subject experiencing HMB, or a subject experiencing AUB. In some embodiments, a threshold amount of a cell type is an upper threshold, such as the maximum 2020373103   25 Mar 2026 amount of a cell type measured during normal menstrual bleeding. In some embodiments, a threshold amount of a cell type is a lower threshold, such as the minimum amount of a cell type measured during normal menstrual bleeding. In some embodiments, a threshold of an amount of a cell type is determined by measuring an amount of a cell type of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing normal menstrual bleeding. In some embodiments, a threshold of an amount of a cell type is determined by measuring the amount of a cell type of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing HMB. In some embodiments, a threshold of an amount of a cell type is determined by measuring the amount of a cell type of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing AUB. In some embodiments, a threshold of an amount of a cell type is determined in a sample of fluid collected from a vaginal cavity other than menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects. In some embodiments, a threshold of an amount of a cell type is determined by comparing the amount of a cell type of a sample of menstrual fluid of subjects experiencing normal menstrual bleeding and HMB or subjects experiencing normal menstrual bleeding and AUB. In some embodiments, a threshold of an amount of a cell type is an average amount of a cell type, a mean amount of a cell type, a mode gene expression, a maximum gene expression, a minimum gene expression, an average gene expression plus 1, 2, or 3 standard deviations of gene expression, or an average gene expression minus 1, 2, or 3 standard deviations of gene expression.

[0113] In some embodiments, an amount of a cell type is increased compared to the normal amount of that cell type or compared with a predetermined threshold. In some embodiments, the amount of a cell type is increased by at least about 10%, 50%, 100%, 150%, or 200%.

[0114] In some embodiments, an amount of a cell type is decreased compared to the normal amount of that cell type or compared with a predetermined threshold. In some embodiments, an amount of a cell type is decreased by at least about 10%, 25%, 50%, 75%, or 90%.

[0115] In some embodiments, a ratio of cell types is increased compared to the normal amount of that cell type or compared with a predetermined threshold. In some embodiments, a ratio of cell types is increased by at least about 10%, 25%, 50%, 75%, or 90%. In some embodiments, a ratio of cell types is decreased compared to the normal amount of that cell type or compared with a predetermined threshold. In some embodiments, a ratio of cell types is decreased by at least about 10%, 25%, 50%, 75%, or 90%.

[0116] In some embodiments, a difference in one or more types of cells found in a fluid sample collected from a vaginal cavity such as menstrual fluid is indicative of HMB, AUB, a menstrual 2020373103   25 Mar 2026 cycle disorder, or another disorder. In some embodiments, a presence of one or more types of cells found in menstrual fluid is indicative of HMB, AUB, a menstrual cycle disorder, or another disorder. Nucleic Acid Content

[0117] In some embodiments of methods herein, fluid from the vaginal cavity of a subject, such as menstrual blood, comprises a nucleic acid content, which is a total nucleic acid content. In some embodiments, a nucleic acid content (e.g., total nucleic acid) is used as a proxy for loss of material during menstruation. In some embodiments, loss of material describes cells or tissue which is lost during menstruation. In some embodiments, material lost comprises that of the subject, of a fetus, of a bacterium, of a virus, or of a fungus. In some embodiments, higher nucleic acid content in menstrual fluid indicates a higher loss of material, while lower nucleic acid content in menstrual fluid indicates a lower loss of material. During HMB or AUB, nucleic acid content is altered when compared with that of normal menstrual bleeding, indicating that during HMB or AUB, the amount of material lost through menstruation is different than during normal menstrual bleeding.

[0118] In some methods, a nucleic acid is measured from a fluid sample obtained from the vaginal cavity of a subject such as menstrual fluid. In some embodiments, nucleic acid is nucleic acid which is present in a collected sample of menstrual fluid from a subject. In some embodiments, nucleic acid is cellular nucleic acid or free nucleic acid. In some embodiments, nucleic acid is nucleic acid of the subject, or is nucleic acid of a virus, bacteria, or fungus present in the menstrual fluid of the subject.

[0119] In some embodiments, nucleic acid in such fluid sample comprises deoxyribonucleic acid (DNA) or ribonucleic acid (RNA). In some embodiments, DNA measured comprises chromosomal DNA or mitochondrial DNA. In some embodiments, RNA measured comprises rRNA, tRNA, mRNA, siRNA, or snRNA. In some embodiments, nucleic acid measured comprises maternal nucleic acid or fetal nucleic acid.

[0120] In some embodiments, total amount of nucleic acid, total DNA, or total RNA is measured. In some embodiments, a ratio of DNA and total nucleic acid, a ratio of DNA and RNA, or a ratio of RNA and total nucleic acid is measured or calculated from a menstrual fluid sample. In some embodiments, a ratio of a type of RNA to total RNA or total nucleic acid (e.g., mRNA to total RNA or mRNA to total nucleic acid) is measured or calculated from a menstrual fluid sample. In some embodiments, a ratio of a type of DNA to total DNA or total nucleic acid (e.g., mitochondrial DNA to total DNA or mitochondrial DNA to total nucleic acid) is measured 2020373103   25 Mar 2026 or calculated from a menstrual fluid sample. In some embodiments, an amount or ratio of nucleic acid, DNA, or RNA is compared with an amount or ratio of nucleic acid, DNA, or RNA from menstrual fluid of a female with normal menstrual bleeding, from menstrual fluid of a female with HMB, or from menstrual fluid of a female with AUB.

[0121] In some embodiments, prior to quantification, nucleic acid, RNA, or DNA is extracted, removed, or purified from a sample for measurement. In some embodiments, extraction methods include organic extraction, Chelex extraction, and solid phase extraction. In some embodiments, organic extraction comprises the addition of one or more chemical solutions to a menstrual fluid sample, lysis, an extraction (e.g., a phenol chloroform extraction), and precipitation (e.g., ethanol precipitation). In some embodiments, Chelex extraction comprises adding a Chelex resin to a menstrual fluid sample, boiling, vortexing, centrifuging, and collecting a supernatant comprising DNA. In some embodiments, solid phase extraction comprises exploiting the ability of DNA to bind to silica by encouraging the binding of DNA to silica (e.g., a column comprising silica or beads comprising silica), perhaps using one or more chaotropic salts.

[0122] In some embodiments, extraction, removal, or purification of nucleic acid, RNA, or DNA comprises removal of protein, or other cellular components from a fraction comprising nucleic acid, RNA, or DNA. In some embodiments, extraction, removal, or purification of nucleic acid, RNA, or DNA comprises precipitating and subsequently re-dissolving the nucleic acid, RNA, or DNA. In some embodiments, gravity filtration, vacuum filtration, or centrifugal filtration is employed for extraction, removal, or purification of nucleic acid, RNA, or DNA, perhaps followed by an elution step. In some embodiments, nucleic acid, RNA, or DNA is gel-purified or purified using a filter paper based lysis and elution method.

[0123] In some embodiments, nucleic acid from a sample is sequenced using an acceptable sequencing method. In some embodiments, sequencing is performed on the total nucleic acid, or on a subset of the nucleic acid, such as RNA, rRNA, tRNA, mRNA, siRNA, snRNA, DNA, chromosomal DNA, or mitochondrial DNA. Nucleic acid sequenced is nucleic acid of the subject, nucleic acid of a microbiome of the subject (e.g., including viral, bacterial, and / or fungal nucleic acid), or a combination thereof. In some embodiments, basic sequencing methods include Maxam-Gilbert sequencing or chain termination methods. In some embodiments, advanced and de novo sequencing methods includes shotgun sequencing or bridge PCR. In some embodiments, high throughput sequencing methods includes massively parallel signature sequencing, Polony sequencing, 454 pyrosequencing, Illumina sequencing, Combinatorial probe anchor synthesis, SOLiD sequencing, Ion Torrent semiconductor sequencing, DNA nanoball 2020373103   25 Mar 2026 sequencing, Heliscope single molecule sequencing, single molecule real time sequencing, nanopore DNA sequencing, or sequencing using microfluidic systems. In some embodiments, sequencing techniques includes tunneling currents DNA sequencing, sequencing by hybridization, sequencing with mass spectrometry, microfluidic Sanger sequencing, microscopybased sequencing, RNAP sequencing, or in vitro virus high-throughput sequencing.

[0124] In some embodiments, nucleic acid in a sample, including total nucleic acid, DNA, or RNA, is quantified using an acceptable method. In some embodiments, a common method for quantifying nucleic acid, DNA, or RNA is spectrophotometric analysis. Briefly, a spectrophotometer is used to determine the average concentration and purity of DNA, RNA, or both in a sample, by exploiting the light absorbing properties of nucleic acids. In some embodiments, nucleic acids absorb ultraviolet light in a specific pattern. In some embodiments, a spectrophotometer exposes a sample comprising DNA, RNA, or both to ultraviolet light, which has a wavelength of about 260 nm, and measure the light which passes through the sample. In some cases, optical density of the sample is calculated from the intensity of the light which is transmitted through the sample and detected using Beer’s law: 0D = Log g) , wherein OD is the optical density of a sample, Ii is the intensity of the incident light (e.g., the light applied to a sample), and It is the transmitted light (e.g., the light detected after passing through a sample). Concentration of nucleic acid in a sample is calculated from optical density as: C = 50 gg / mL x DD x d / , wherein C is the concentration of the DNA or RNA in the sample, OD is the optical density (e.g., as calculated by Beer’s law), and df is a dilution factor of the sample.

[0125] In some embodiments, another common method for quantifying nucleic acid, DNA, or RNA is fluorescence tagging, e.g., in the presence of a nucleotide dye. Briefly, DNA or RNA is tagged with a fluorescent tag, which is a fluorescent nucleotide dye, and the nucleotide dye that binds to the RNA or DNA selectively fluoresces when bound. In some embodiments, dyes is UV fluorescent dyes. In some embodiments, a tagging method is modified to use a non-fluorescent nucleotide dye, such as a visual dye. In some embodiments, DNA dyes include ethidium bromide, SYBR gold, SYBR green, SYBR safe, Eva green, propidium iodide, crystal violet, dUTP-conjugated probes, 4’,6-diamidino-2-phenylindole, 7-aminoactinomycin D, Hoechst 33258 (33342, 34580), or YOYO-1 / DiYO-1 / TOTO-1 / DiTO-1. 2020373103   25 Mar 2026

[0126] In some embodiments, nucleic acid is measured or quantified on any day a subject is experiencing menstrual bleeding. In some embodiments, nucleic acid is measured more than once during menstrual bleeding. In some embodiments, nucleic acid is measured on day 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 of menstrual bleeding. In some embodiments, nucleic acid is measured after 30 days of menstrual bleeding. In some embodiments, nucleic acid is measured or quantified when a subject is not experiencing menstrual bleeding, such as between two menstrual windows.

[0127] In some embodiments, nucleic acid in a sample, including total nucleic acid, DNA, RNA, or a ratio or metric thereof, is increased or decreased compared with a predetermined threshold. In some embodiments, a predetermined threshold of nucleic acid is determined from a reference sample, such as a fluid collected from the vaginal cavity of a control subject, such as menstrual fluid collected from a subject experiencing normal menstrual bleeding, a subject experiencing HMB, or a subject experiencing AUB. In some embodiments, a threshold of nucleic acid is an upper threshold, such as the maximum nucleic acid measured during normal menstrual bleeding. In some embodiments, a threshold of nucleic acid is a lower threshold, such as the minimum nucleic acid measured during normal menstrual bleeding. In some embodiments, a threshold of nucleic acid is determined by measuring the nucleic acid of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing normal menstrual bleeding. In some embodiments, a threshold of nucleic acid is determined by measuring the nucleic acid of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing HMB. In some embodiments, a threshold of nucleic acid is determined by measuring the nucleic acid of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing AUB. In some embodiments, a threshold of nucleic acid is determined in a sample of fluid collected from a vaginal cavity other than menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects. In some embodiments, a threshold of nucleic acid is determined by comparing the nucleic acid of a sample of menstrual fluid of subjects experiencing normal menstrual bleeding and HMB, or subjects experiencing normal menstrual bleeding and AUB. In some embodiments, a threshold of nucleic acid is an average amount of nucleic acid, a mean amount of nucleic acid, a mode amount of nucleic acid, a maximum amount of nucleic acid, a minimum amount of nucleic acid, an average amount of nucleic acid plus 1, 2, or 3 standard deviations of an amount of nucleic acid, or an average amount of nucleic acid minus 1, 2, or 3 standard deviations of an amount of nucleic acid. 2020373103   25 Mar 2026

[0128] In some embodiments, an increase in nucleic acid is an increase of at least about 1%, about 5%, about 10%, about 15%, about 25%, about 25%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 100%, about 150%, about 200%, or more compared with a predetermined threshold. In some embodiments, a decrease in nucleic acid is a decrease of at least about 1%, about 5%, about 10%, about 15%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 100% compared with a predetermined threshold. In some embodiments, an increase in nucleic acid in a sample is indicative of HMB, AUB, a menstrual cycle disorder, or another disorder. In some embodiments, decrease in nucleic acid in a sample is indicative of HMB, AUB, a menstrual cycle disorder, or another disorder. Biomarkers

[0129] In some embodiments of methods herein, a fluid sample from the vaginal cavity of a subject comprises a biological marker or biomarker profile. In some embodiments, during HMB or AUB, this biomarker profile is altered. In some embodiments, biomarkers is of a tissue or cell, such as a fallopian, ovarian, uterine, endometrial, cervical, or vaginal tissue or cell that is found in a menstrual fluid sample of a subject. In some embodiments, a biomarker is of a component of blood, such as an erythrocyte, plasma, a leukocyte, a platelet, or other component. In some embodiments, a biomarker is common to any set of subjects or a set of all subjects. In some embodiments, a biomarker is common to a set of subjects experiencing normal menstrual bleeding, HMB, or AUB. In some embodiments, a biomarker is unique to a set of subjects experiencing normal menstrual bleeding, HMB, or AUB. One or biomarkers of a menstrual fluid sample is indicative of HMB, AUB, a menstrual cycle disease or disorder, or another disease or disorder.

[0130] In some embodiments, a biomarker is measured on any day a subject is experiencing menstrual bleeding. In some embodiments, a biomarker is measured more than once during menstrual bleeding. Flow rate is measured on day 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 of menstrual bleeding. In some embodiments, a biomarker is measured between days 1 and 30, between days 5 and 30, between days 10 and 30, between days 20 and 30, between days 1 and 20, between days 5 and 20, between days 10 and 20, between days 1 and 10, between days 5 and 10, between days 1 and 7, between days 1 and 6, between days 1 and 5, between days 1 and 4, between days 1 and 3, between days 1 and 2, between days 2 and 7, between days 2 and 6, between days 2 and 5, 2020373103   25 Mar 2026 between days 2 and 4, between days 2 and 3, between days 3 and 7, between days 3 and 6, between days 3 and 5, between days 3 and 4, between days 4 and 7, between days 4 and 6, between days 4 and 5, between days 5 and 7, between days 5 and 6, or between days 6 and 7 of menstrual bleeding. Sometimes, a biomarker is measured after 30 days of menstrual bleeding. In some embodiments, a biomarker is measured 1, 2, 3, 4, 5, or more times during a menstrual cycle.

[0131] In some embodiments, one or more biomarkers is assayed in a sample collected when a subject is not menstruating, such as between two menstrual windows, such as any time a sample may be collected as described herein. In some embodiments, a biomarker assayed when a subject is not menstruating is a biomarker that is measured when a subject is menstruating, or a biomarker that cannot be measured when a subject is menstruating. In some embodiments, a biomarker measured from a vaginal fluid sample outside the menstrual window is indicative of a disorder. In some embodiments, a biomarker measured from a vaginal sample outside the menstrual window is used in combination with a biomarker measured from a menstrual fluid sample to indicate a menstrual cycle disorder, HMB, or AUB.

[0132] In some methods, one or more biomarkers in a sample collected from a vaginal cavity such as menstrual fluid is detected or measured. In some embodiments, biomarkers comprise protein biomarkers or gene target biomarkers. In some embodiments, a protein biomarker is a protein which is found in menstrual blood. In some embodiments, a protein biomarker is either present or absent in normal menstrual blood or in menstrual blood from a subject experiencing HMB or AUB. In some embodiments, a presence or absence of a protein biomarker in a sample of menstrual blood is indicative of a menstrual cycle disorder.

[0133] In some embodiments, proteomic analysis of a sample collected from a vaginal cavity such as menstrual fluid identify protein biomarkers which is found in a sample such as a menstrual fluid sample. In some embodiments, a protein biomarkers is unique to menstrual fluid. In some embodiments, a protein biomarkers is unique to another fluid collected from a vaginal cavity. In some embodiments, a proteins are not be found in whole blood or other vaginal discharge or fluid. In some embodiments, identification or measurement of such proteins in a sample provides insight into the health, e.g., menstrual health or reproductive health of a female. In some instances, at least 1, 2, 3, 4, 5 6, 7, 8, 9, 10, 50, 100, 200, 300, 400, or 500 protein biomarkers is identified as unique to menstrual fluid or another fluid collected from a vaginal cavity. In some embodiments, a set of about 385 proteins makes up a unique set of protein biomarkers in menstrual fluid or another fluid collected from a vaginal cavity. In some 2020373103   25 Mar 2026 embodiments, the protein biomarkers are integral to the menstrual cycle, at times including the endometrial cycle and is used to assess normal menstrual bleeding, HMB, or AUB. In some embodiments, a list of protein biomarkers which is found in a menstrual fluid sample is presented in Table 1.

[0134] In some embodiments, a protein biomarker is measured using techniques such as enzyme linked immunosorbent assay (ELISA), western blot, LC-MS, flow-cytometry, immunohistochemistry, or other technique capable of detecting or measuring a protein biomarker.

[0135] In some embodiments, an amount of a protein biomarker is measured and is expressed as an absolute amount or a relative amount. In some embodiments, an absolute amount of a protein biomarker is measured as an amount of a biomarker present in a tested sample and is expressed for example as a number of molecules, as a mass (e.g., mg), as a number of molecules per volume, or as a mass per volume in a menstrual fluid sample. In some embodiments, a relative amount of a protein biomarker is measured as an amount of a protein biomarker normalized to another value. In some embodiments, a relative amount of a protein is normalized for example to a total amount of protein or protein biomarker in a menstrual fluid sample, to a total amount of a specific protein, protein biomarker, set of proteins, or set of protein biomarkers in a menstrual fluid sample, to an amount of menstrual fluid in a menstrual fluid sample, to an amount of cells in a menstrual fluid sample, to an amount of a cell type in a menstrual fluid sample, to a gene expression in a menstrual fluid sample, to a gene target biomarker in a menstrual fluid sample, or to an amount of a same protein biomarker in a menstrual fluid sample from a subject experiencing normal menstrual bleeding.

[0136] In some embodiments, a protein biomarker is increased or decreased in a subject experiencing a disorder such as a menstrual cycle disorder, HMB or AUB. In some embodiments, an increase or decrease is determined by comparing the measured level of a protein biomarker to the measured level of a protein biomarker in a reference sample, such as fluid collected from the vaginal cavity of a control subject, such as a sample of menstrual fluid from a reference subject or subject experiencing normal menstrual bleeding.

[0137] In some embodiments, a protein biomarker is increased or decreased compared with a predetermined threshold amount. In some embodiments, a predetermined threshold of a protein biomarker is determined from a reference sample, such as a reference sample collected from a subject having a menstrual cycle disorder, a subject experiencing normal menstrual bleeding, a subject experiencing HMB, or a subject experiencing AUB. In some embodiments, a threshold 2020373103   25 Mar 2026 of a protein biomarker is determined by measuring an amount of a protein biomarker in samples of menstrual blood of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing normal menstrual bleeding. In some embodiments, a threshold value is determined by measuring the amount of a protein biomarker of the menstrual blood of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing HMB. In some embodiments, a threshold value is determined by measuring the amount of a protein biomarker of the menstrual blood of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing AUB. In some embodiments, a threshold of a protein biomarker is determined in a sample of fluid collected from a vaginal cavity other than menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects. In some embodiments, a threshold value is determined by comparing an amount of a protein biomarker in samples of subjects experiencing normal menstrual bleeding and HMB or subjects experiencing normal menstrual bleeding and AUB. In some embodiments, a threshold of a protein biomarker is an average amount of a protein biomarker, a mean amount of a protein biomarker, a mode amount of a protein biomarker, a maximum amount of a protein biomarker, a minimum amount of a protein biomarker, an average amount plus 1, 2, or 3 standard deviations of a protein biomarker, or an average amount minus 1, 2, or 3 standard deviations of a protein biomarker.

[0138] In some embodiments, an amount of a protein biomarker can increase at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 300%, 400%, or 500% compared with a predetermined threshold. In some embodiments, an amount of a protein biomarker can decrease by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%, compared with a predetermined threshold. In some embodiments, an increase or decrease in an amount of a protein biomarker is associated with a disorder such as a menstrual cycle disorder, HMB or AUB. In some embodiments, an increase or decrease in a level of a protein biomarker in a sample of menstrual blood is indicative of a menstrual cycle disorder.

[0139] In some embodiments, relative levels of two or more protein biomarkers is determined. In some embodiments, a ratio of two protein biomarkers is determined and compared with the ratio of the same two protein biomarkers in a reference sample. In some embodiments, an increase or decrease in said ratio is detected in a subject experiencing HMB or AUB. In some embodiments, an increase or decrease in said ratio in a sample of menstrual fluid is indicative of a menstrual cycle disorder.

[0140] In some embodiments, a panel of protein biomarkers is measured. In some embodiments, a panel of protein biomarkers comprises at least 2, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 protein biomarkers. In some embodiments, at least 1, 2, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 2020373103   25 Mar 2026 100 protein biomarkers in the panel is altered (increased or decreased) in a sample collected from a subject experiencing HMB or AUB.

[0141] In some embodiments, one or more markers from a panel of biomarkers is measured in menstrual fluid. In some embodiments, some such biomarkers is measured in a sample of a fluid collected from a vaginal cavity that is not menstrual fluid. In some embodiments, proteins which is biomarkers in menstrual fluid may be found in Table 1. Table 1: Menstrual fluid protein biomarkers Accession # Protein Name Accession # Protein Name P15428 15-hydroxyprostaglandin dehydrogenase [NAD+] Q9H293 Interleukin-25 O15120 "1 -acyl-sn-glycerol-3 -phosphate acyltransferase beta 1-acyl-sn-glycerol-3-phosphate acyltransferase beta" Q96CT2 Kelch-like protein 29 P62979 40S ribosomal protein S27a P02533 Keratin, type I cytoskeletal 14 Q8NHW5 60S acidic ribosome protein P0-like P08727 Keratin, type I cytoskeletal 19 Q96L21 60S ribosomal protein L10-like P08779 Keratin, type I cytoskeletal 16 P83881 60S ribosomal protein L36a Q2M2I5 Keratin, type I cytoskeletal 24 Q969Q0 60S ribosomal protein L36a-like P13645 Keratin, type I cytoskeletal 10 P40429 60S ribosome protein L13A P13646 Keratin, type I cytoskeletal 13 Q9Y3U8 60S ribosome protein L36 P35527 Keratin, type I cytoskeletal 9 O95450 A disintegrin and metalloproteinase with thromb Q9NSB4 Keratin, type II cuticular Hb2 Q9UP79 A disintegrin and metalloproteinase with thromb O95678 Keratin, type II cytoskeletal 75 P07510 Acetylcholine receptor subunit gamma P05787 Keratin, type II cytoskeletal 8 Q9BWD1 Acetyl-CoA acetyltransferase, cyt P08729 Keratin, type II cytoskeletal 7 P68133 Actin, alpha skeletal muscle P12035 Keratin, type II cytoskeletal 3 P63267 Actin, gamma-enteric smooth muscle P19013 Keratin, type II cytoskeletal 4 Q9C0K3 Actin-related protein 11 Q01546 Keratin, type II cytoskeletal 2 oral Q9P1U1 Actin-related protein 3B Q14CN4 Keratin, type II cytoskeletal 72 Q9BZ11 ADAM 33 Q5XKE5 Keratin, type II cytoskeletal 79 Q9H6B4 Adipocyte adhesion molecule Q6KB66 Keratin, type II cytoskeletal 80 P12235 ADP / ATP translocase 1 Q86Y46 Keratin, type II cytoskeletal 73 Q9H0C2 ADP / ATP translocase 4 Q8N1N4 Keratin, type II cytoskeletal 78 P61204 ADP-ribosylation factor 3 P04264 Keratin, type II cytoskeletal 1 P05187 Alkaline ph P35908 Keratin, type II cytoskeletal 2 epidermal Q9BQI0 Allograft inflammatory factor 1 -like P13647 Keratin, type II cytoskeletal 5 P61647 Alpha-2,8-sialyltransferase 8F P02538 Keratin, type II cytoskeletal 6A P35609 Alpha-actinin-2 P04259 Keratin, type II cytoskeletal 6B Q6S8J3 ANKRD26-like family C member 1A P48668 Keratin, type II cytoskeletal 6C A5A3E0 ANKRD26-like family C member 1B Q5T7P3 Late cornified envelope protein 1B Q5VYY1 Ankyrin repeat domain-containing protein 22 Q5TA79 Late cornified envelope protein 2A Q5VT79 Annexin A8-like protein 2 O14633 Late cornified envelope protein 2B Q16671 Anti-Muellerian hormone type-2 receptor Q5TA81 Late cornified envelope protein 2C P53680 AP-2 complex subunit sigma-1 Q5TA82 Late cornified envelope protein 2D P02654 Apolipoprotein C-I Q5TA76 Late cornified envelope protein 3A P08519 Apolipoprotein(a) Q9BYE3 Late cornified envelope protein 3D 2020373103   25 Mar 2026 P20292 Arachidonate 5 -lipoxygenase-activating protein Q5TA78 Late cornified envelope protein 4A P15848 Arylsulfatase B Q2I0M4 Leucine-rich repeat-containing protein 26 Q15121 Astrocytic ph Q8N423 Leukocyte immunoglobulin-like receptor subfamily B member 2 Q8IZY2 ATP-binding cassette sub-family A member 7 O75197 Low-density lipoprotein receptor-related protein 5 O00571 ATP-dependent RNA helicase DDX3X Q14114 Low-density lipoprotein receptor-related protein 8 O15523 ATP-dependent RNA helicase DDX3Y P01229 Lutropin subunit beta P16066 Atrial natriuretic peptide receptor A P07333 Macrophage colony-stimulating factor 1 receptor Q8NFJ9 Bardet-Biedl syndrome 1 protein Q9ULC4 Ma lignant T call amplified sequence 1 Q9BYX7 Beta-actin-like protein 3 Q8WXG6 MAP kinase-activating death domain protein P35612 Beta-adducin Q495T6 Membrane metallo-endopeptidase-like 1 Q9Y2P5 Bile acyl-CoA synthetase Q15049 Membrane protein MLC1 O14514 Brain-specific angiogenesis inhibitor 1 Q14696 Mesoderm development candidate 2 Q8WY22 BRI3-binding protein P10620 Microsomal glutathione S-transferase 1 Q6PL45 BRICH Q96DA6 Mitochondrial import inner membrane translocase subunit TIM14 O95696 Bromodomain-containing protein 1 P27361 Mitogen-activated protein kinase 3 Q8WVV5 Butyrophilin subfamily 2 member A2 O14950 Myosin regulatory light chain 12B Q9HCU4 Cadherin EGF LAG seven-pass G-type receptor 2 Q76KP1 N-acetyl-beta-gluc Q9UJ99 Cadherin-22 P51970 NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 8 Q9Y6N3 Calcium-activated chloride channel regulator family member 3 Q96PH1 NADPH oxidase 5 P07451 Carbonic anhydrase 3 O95944 Natural cytotoxicity triggering receptor 2 P49747 Cartilage oligomeric matrix protein Q9P2S2 Neurexin-2-alpha Q8N126 Cell adhesion molecule 3 P58401 Neurexin-2-beta O95833 Chloride intracellular channel protein 3 Q99466 Neurogenic locus notch homolog protein 4 P11597 Cholesteryl ester transfer protein P48645 Neuromedin-U P01233 Choriogonadotropin subunit beta Q8N729 Neuropeptide W A6NKQ9 Choriogonadotropin subunit beta variant 1 P12838 Neutrophil defensin 4 Q6NT52 Choriogonadotropin subunit beta variant 2 O15381 Nuclear val Q9P2D1 Chromodomain-helicase-DNA-binding protein 7 Q8NH81 Olfactory receptor 10G6 Q9Y281 Cofilin-2 O43612 Orexin Q8N6L0 Coiled-coil domain-containing protein 155 Q9HC10 Otoferlin A1A5D9 Coiled-coil domain-containing protein 64B P50542 Peroxisomal targeting signal 1 receptor Q14050 Collagen alpha-3(IX) chain Q9BTY2 Plasma alpha-L-fuc Q96CG8 Collagen triple helix repeat-containing protein 1 Q9UIW2 Plexin-A1 Q9NPY3 Complement component C1q receptor Q9Y4D7 Plexin-D1 Q9HCH3 Copine-5 Q7Z5L7 Podocan O95741 Copine-6 Q6PEZ8 Podocan-like protein 1 Q9UBL6 Copine-7 Q92692 Poliovirus receptor-related protein 2 2020373103   25 Mar 2026 Q86YQ8 Copine-8 Q96NY8 Poliovirus receptor-related protein 4 Q9P0M6 Core histone macro-H2A.2 O15354 Probable G-protein coupled receptor 37 Q9BYD5 Cornifelin Q8N0Y7 Probable phosphoglycerate mutase 4 Q9H2A7 C-X-C motif chemokine 16 O43555 Progonadoliberin-2 O14949 Cytochrome b-c1 complex subunit 8 Q9HC23 Prokineticin-2 P09669 Cytochrome c oxidase subunit 6C Q8IVL6 Prolyl 3 -hydroxylase 3 Q02928 Cytochrome P450 4A11 Q16647 Prostacyclin synthase Q5TCH4 Cytochrome P450 4A22 Q8TAA3 Proteasome subunit alpha type-7-like Q14008 Cytoskeleton-associated protein 5 Q86UD0 Protein C9orf140 Q96GG9 DCN1 -like protein 1 Q9BT09 Protein canopy homolog 3 Q9BTC0 Death-inducer obliterator 1 P05129 Protein kinase C gamma type Q08495 Dematin Q8N114 Protein shisa-5 Q8TEH3 DENN domain-containing protein 1A Q93097 Protein Wnt-2b Q8IV53 DENN domain-containing protein 1C Q9Y6F9 Protein Wnt-6 Q9H4A9 Dipeptidase 2 Q9UM07 Protein-arginine deiminase type-4 Q9H4B8 Dipeptidase 3 Q9UN74 Protocadherin alpha-4 O95886 Disks large-associated protein 3 Q9Y5H6 Protocadherin alpha-8 O95147 Dual specificity protein ph Q9Y5I4 Protocadherin alpha-C2 Q9P225 Dynein heavy chain 2, axonemal Q14517 Protocadherin Fat 1 Q96JB1 Dynein heavy chain 8, axonemal Q96JQ0 Protocadherin-16 Q5XPI4 E3 ubiquitin-protein ligase RNF123 Q2VWP7 Protogenin P84090 Enhancer of rudimentary homolog P07949 Proto-oncogene tyr P52797 Ephrin-A3 Q5JNZ5 Putative 40S rib P52798 Ephrin-A4 A6NMY6 Putative annexin A2-like protein Q08345 Epithelial discoidin domain-containing receptor 1 A8MZA4 Putative golgin subfamily A member 6-like protein 6 Q9NZJ5 Eukaryotic translation initiation factor 2-alpha kinase 3 P48741 Putative heat shock 70 kDa protein 7 Q9UKV8 Eukaryotic translation initiation factor 2C 2 Q58FF7 Putative heat shock protein HSP 90-beta-3 Q16658 Fascin Q6DN03 Putative histone H2B type 2-C Q9Y3I1 F-box only protein 7 Q6DRA6 Putative histone H2B type 2-D Q9NYL4 FK506-binding protein 11 Q92928 Putative Ras-related protein Rab-1C Q5SZK8 FRAS1-related extracellular matrix protein 2 Q5VYJ4 Putative small nuclear ribonucleoprotein polypeptide E-like protein 1 O15552 Free fatty acid receptor 2 A6NGW2 Putative stereocilin-like protein Q9H479 Fructosamine-3-kinase Q9H853 Putative tubulin-like protein alpha-4B O75899 Gamma-aminobutyric acid type B receptor subunit 2 A6NJS3 Putative V-set and immunoglobulin domain-containing-like protein ENSP00000303034 Q9GZZ7 GDNF family receptor alpha-4 P35241 Radixin Q8N2G8 GH3 domain-containing protein A6NIZ1 Ras-related protein Rap-1b-like protein P47871 Glucagon receptor Q92932 Receptor-type tyr O76003 Glutaredoxin-3 Q9NZ71 Regulator of telomere elongation helicase 1 P08263 Glutathione S-transferase A1 Q96D15 Reticulocalbin-3 P09210 Glutathione S-transferase A2 Q02846 Retinal guanylyl cyclase 1 Q16772 Glutathione S-transferase A3 P11908 Ribose-phosphate pyrophosphokinase 2 Q9NZD2 Glycolipid transfer protein P21108 Ribose-phosphate pyrophosphokinase 3 P02724 Glycophorin-A Q15050 Ribosome biogenesis regulatory protein homolog 2020373103   25 Mar 2026 Q8N3Y3 Glycosyltransferase-like protein LARGE2 O94885 SAM and SH3 domain-containing protein 1 Q8N7Z2 Golgin subfamily A member 6-like protein 1 P10523 S-arrestin Q96HH9 GRAM domain-containing protein 3 Q96GP6 Scavenger receptor class F member 2 O15496 Group 10 secretory ph P09683 Secretin P01116 GTPase KRas Q13018 Secretory ph Q9UBI6 Guanine nucleotide-binding protein G(I) / G(S) / G(O) subunit gamma-12 Q9NPR2 Semaphorin-4B Q9HAV0 Guanine nucleotide-binding protein subunit beta-4 Q9H3T3 Semaphorin-6B Q6B0K9 Hemoglobin subunit mu Q9H3T2 Semaphorin-6C P09105 Hemoglobin subunit theta-1 Q9UQ35 Serine / arginine repetitive matrix protein 2 P02008 Hemoglobin subunit zeta Q9BZL6 Serine / threonine-protein kinase D2 Q8TDQ0 Hepatitis A virus cellular receptor 2 Q9UL54 Serine / threonine-protein kinase TAO2 Q04756 Hepatocyte growth factor activator Q8IYT8 Serine / threonine-protein kinase ULK2 P81172 Hepcidin P30154 Serine / threonine-protein ph P55795 Heterogeneous nuclear ribonucleoprotein H2 P36873 Serine / threonine-protein ph Q6UXD1 Histidine-rich carboxyl terminus protein 1 P62714 Serine / threonine-protein ph Q92522 Histone H1x Q5TCZ1 SH3 and PX domain-containing protein 2A P04908 Histone H2A type 1-B / E Q96PQ1 Sialic acid-binding Ig-like lectin 12 P20671 Histone H2A type 1 -D Q9NYZ4 Sialic acid-binding Ig-like lectin 8 Q96KK5 Histone H2A type 1 -H Q9NQ36 Signal peptide, CUB and EGF-like domain-containing protein 2 Q99878 Histone H2A type 1 -J P49771 SL cytokine Q6FI13 Histone H2A type 2-A Q96A28 SLAM family member 9 Q16777 Histone H2A type 2-C Q8N130 Sodium-dependent phosphate transport protein 2C Q9BTM1 Histone H2A.J Q9UMY4 Sorting nexin-12 P16104 Histone H2A.x Q7RTU9 Stereocilin P58876 Histone H2B type 1-D P78539 Sushi repeat-containing protein SRPX Q93079 Histone H2B type 1-H Q7RTX1 Taste receptor type 1 member 1 P06899 Histone H2B type 1-J Q7RTX0 Taste receptor type 1 member 3 Q99879 Histone H2B type 1-M Q9BZG2 Testicular acid ph P23527 Histone H2B type 1-O Q9UI38 Testis-specific protease-like protein 50 Q16778 Histone H2B type 2-E Q9UPZ6 Thrombospondin type-1 domaincontaining protein 7A Q5QNW6 Histone H2B type 2-F P01266 Thyroglobulin P57053 Histone H2B type F-S Q9UPV9 Trafficking kinesin-binding protein 1 P84243 Histone H3.3 Q969E4 Transcription elongation factor A protein-like 3 Q6NXT2 Histone H3-like Q6IPX3 Transcription elongation factor A protein-like 6 Q99525 Histone H4-like protein type G P30536 Translocator protein P78426 Homeobox protein Nkx-6.1 P43307 Translocon-associated protein subunit alpha P01742 Ig heavy chain V-I region EU Q9BVT8 Transmembrane and ubiquitin-like domain-containing protein 1 P06326 Ig heavy chain V-I region Mot Q9Y3Q3 Transmembrane emp24 domaincontaining protein 3 P01761 Ig heavy chain V-I region SIE A6NL71 Transmembrane protease, serine 11E2 P01760 Ig heavy chain V-I region WOL Q9UK28 Transmembrane protein 59-like P01777 Ig heavy chain V-III region TEI A6NKL6 Transmembrane protein TTMA 2020373103   25 Mar 2026 P01765 Ig heavy chain V-III region TIL Q13428 Treacle protein P01763 Ig heavy chain V-III region WEA Q9HCJ0 Trinucleotide repeat-containing gene 6C protein P01594 Ig kappa chain V-I region AU P28289 Tropomodulin-1 P04432 Ig kappa chain V-I region Daudi P07477 Trypsin-1 P01598 Ig kappa chain V-I region EU P07478 Trypsin-2 P01599 Ig kappa chain V-I region Gal P35030 Trypsin-3 P01600 Ig kappa chain V-I region Hau Q96A98 Tuberoinfundibular peptide of 39 residues P01601 Ig kappa chain V-I region HK101 (Fragment) Q13748 Tubulin alpha-3C / D chain P01607 Ig kappa chain V-I region Rei Q6PEY2 Tubulin alpha-3E chain P01608 Ig kappa chain V-I region Roy Q9NY65 Tubulin alpha-8 chain P04431 Ig kappa chain V-I region Walker Q13885 Tubulin beta-2A chain P80362 Ig kappa chain V-I region WAT Q9BVA1 Tubulin beta-2B chain P01610 Ig kappa chain V-I region WEA Q13509 Tubulin beta-3 chain P06309 Ig kappa chain V-II region GM607 (Fragment) P08138 Tumor necr P06310 Ig kappa chain V-II region RPMI 6410 P41273 Tumor necrosis factor ligand superfamily member 9 P01619 Ig kappa chain V-III region B6 Q13454 Tumor suppressor candidate 3 P04206 Ig kappa chain V-III region GOL Q12792 Twinfilin-1 P01621 Ig kappa chain V-III region NG9 (Fragment) Q9Y4E8 Ubiquitin carboxyl-terminal hydrolase 15 P01624 Ig kappa chain V-III region POM Q8TEY7 Ubiquitin carboxyl-terminal hydrolase 33 P04434 Ig kappa chain V-III region VH (Fragment) Q5VVQ6 Ubiquitin thioesterase OTU1 P01623 Ig kappa chain V-III region WOL P51668 Ubiquitin-conjugating enzyme E2 D1 P06314 Ig kappa chain V-IV region B17 P61077 Ubiquitin-conjugating enzyme E2 D3 P06313 Ig kappa chain V-IV region JI Q9Y2X8 Ubiquitin-conjugating enzyme E2 D4 P06316 Ig lambda chain V-I region BL2 P62253 Ubiquitin-conjugating enzyme E2 G1 P01700 Ig lambda chain V-I region HA Q14376 UDP-glucose 4-epimerase P01699 Ig lambda chain V-I region VOR Q6UX73 Uncharacterized protein C16orf89 P01715 Ig lambda chain V-IV region Bau Q6ICL3 Uncharacterized protein C22orf25 O95373 Importin-7 Q08AI8 Uncharacterized protein C2orf54 Q8WX77 Insulin-like growth factor-binding proteinlike 1 Q5BLP8 Uncharacterized protein C4orf48 P19827 Inter-alpha-trypsin inhibitor heavy chain H1 Q5TF21 Uncharacterized protein C6orf174 P19823 Inter-alpha-trypsin inhibitor heavy chain H2 Q9Y2F5 Uncharacterized protein KIAA0947 Q06033 Inter-alpha-trypsin inhibitor heavy chain H3 Q6ZSJ9 UPF0626 protein B Q9UHA7 Interleukin-1 family member 6 Q969E3 Urocortin-3 Q13261 Interleukin-15 receptor subunit alpha P10746 Uroporphyrinogen-III synthase Q9HBE5 Interleukin-21 receptor P54725 UV excision repair protein RAD23 homolog A Q5T200 Zinc finger CCCH domain-containing protein 13 Q9H0V9 VIP36-like protein Q12836 Zona pellucida sperm-binding protein 4 Q8TEU8 WAP, kazal, immunoglobulin, kunitz and NTR domain-containing protein 2 Q9HCY8 Protein S100-A14

[0142] In some embodiments, a gene target biomarker is a gene target which is present in a sample collected from a vaginal cavity such as menstrual fluid. In some embodiments, a gene target biomarker is either present or absent in a fluid collected from a vaginal cavity, normal 2020373103   25 Mar 2026 menstrual fluid, or in menstrual fluid from a subject experiencing HMB or AUB. In some embodiments, a presence, absence, or value of a gene target biomarker in a sample collected from a vaginal cavity menstrual fluid is indicative of a menstrual cycle disorder.

[0143] In some embodiments, a gene target biomarker in a sample collected from a vaginal cavity such as menstrual fluid is measured using an acceptable method In some embodiments, methods for measuring gene expression include polymerase chain reaction (PCR), quantitative real-time (q-RT-PCR), isothermal PCR, restriction enzyme analysis, RNA sequencing (e.g., sequencing mRNA), northern blotting, serial analysis of gene expression (SAGE), in situ hybridization (ISH), fluorescence ISH (FISH), and microarray analysis, and microarray analysis.

[0144] In some embodiments, an amount of a gene target biomarker is measured, and is expressed as an absolute amount or a relative amount. In some embodiments, absolute expression is measured as an amount of a gene target biomarker present in a tested sample, and is expressed for example as a number of copies, as a mass (e.g., mg), as a number of copies per volume, or as a mass per volume in a menstrual fluid sample. In some embodiments, relative expression is measured as an amount of a gene target biomarker normalized to another value. In some embodiments, relative expression is normalized for example to a total amount of one or more gene target biomarkers in a sample collected from a vaginal cavity such as a menstrual fluid sample, an amount of menstrual fluid in a menstrual fluid sample, an amount of cells in a menstrual fluid sample, an amount of a cell type in a menstrual fluid sample, to an amount of a protein in a menstrual fluid sample, or to an amount of a same gene target biomarker in a menstrual fluid sample from a subject experiencing normal menstrual bleeding.

[0145] In some embodiments, a gene target biomarker is increased or decreased in a subject experiencing a menstrual cycle disorder, HMB, or AUB. In some embodiments, an increase or decrease is determined by comparing the measured level of a gene target biomarker to the measured level of a protein biomarker in a reference sample fluid collected from the vaginal cavity of a control subject, such as a sample of menstrual fluid from a reference subject or subject experiencing normal menstrual bleeding.

[0146] In some embodiments, a gene target biomarker is increased or decreased compared with a predetermined threshold. In some embodiments, a predetermined threshold of a gene target biomarker is determined from a reference sample, such as a reference sample collected from a subject experiencing normal menstrual bleeding, a subject experiencing HMB, or a subject experiencing AUB. In some embodiments, a threshold gene target biomarker is an upper threshold, such as the maximum gene expression measured during normal menstrual bleeding. 2020373103   25 Mar 2026 In some embodiments, a threshold gene target biomarker is a lower threshold, such as the minimum gene expression measured during normal menstrual bleeding. In some embodiments, a threshold of a gene target biomarker is determined by measuring a gene target biomarker of a sample of a fluid collected from a vaginal cavity such as menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing normal menstrual bleeding. In some embodiments, a threshold of gene target biomarker is determined by measuring the gene expression of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing HMB. In some embodiments, a threshold of a gene target biomarker is determined by measuring the gene expression of a sample of menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing AUB. In some embodiments, a threshold of a gene target biomarker is determined in a sample of fluid collected from a vaginal cavity other than menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects. In some embodiments, a threshold of a gene target biomarker is determined by comparing the gene expression of a sample of menstrual fluid of subjects experiencing normal menstrual bleeding and HMB or subjects experiencing normal menstrual bleeding and AUB. In some embodiments, a threshold of a gene target biomarker is an average amount of a gene target biomarker, a mean amount of a gene target biomarker, a mode amount of a gene target biomarker, a maximum amount of a gene target biomarker, a minimum amount of a gene target biomarker, an average amount of a gene target biomarker, plus 1, 2, or 3 standard deviations of gene expression, or an average amount of a gene target biomarker, minus 1, 2, or 3 standard deviations of gene expression.

[0147] In some embodiments, an amount of a gene target biomarker can increase at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 300%, 400%, or 500% compared with a predetermined threshold. In some embodiments, an amount of a gene target biomarker can decrease by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared with a predetermined threshold. In some embodiments, an increase or decrease in an amount of a protein biomarker is associated with HMB or AUB. In some embodiments, an increase or decrease in a level of a protein biomarker in a sample of menstrual fluid is indicative of a menstrual cycle disorder.

[0148] In some embodiments, relative levels of two or more gene target biomarkers is determined. In some embodiments, a ratio of two gene target biomarkers is determined and compared with the ratio of the same two gene target biomarkers in a reference sample. In some embodiments, an increase or decrease in said ratio is detected in a subject experiencing HMB or 2020373103   25 Mar 2026 AUB. In some embodiments, an increase or decrease in said ratio in a sample of menstrual fluid is indicative of a menstrual cycle disorder.

[0149] In some embodiments, a panel of gene target biomarkers is measured. In some embodiments, a panel of gene target biomarkers comprises at least 2, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 gene target biomarkers. In some embodiments, at least 1, 2, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 gene target biomarkers in the panel is altered (increased or decreased) in a sample collected from a subject experiencing HMB or AUB.

[0150] In some embodiments, one or more genes is used as target biomarkers for a menstrual fluid sample from a subject. In some embodiments, gene targets which is biomarkers in menstrual fluid may be found in Table 2. Table 2: Menstrual blood gene target biomarkers Gene Name GNB1 MPZL2 KRT13 RPS21 DEK RPL27A ARL6IP1 RPL22 MLL KRT16 PPDPF C6orf62 IPO7 SCNN1B ENO1 RPS25 EIF1 PTK6 FAM65B EIF4G2 IL4R PLOD1 ARHGEF12 PTRF DNAJC5 HIST1H2AC COPB1 MVP EPHA2 HSPA8 RPL27 MIR3648 HLA-A RPS13 CORO1A CDC42 ETS1 SLC25A39 MIR3687 HLA-E LDHA PRSS8 CLIC4 APLP2 ARHGAP27 NRIP1 TUBB HIPK3 CYLD MED18 CCND2 CDC27 APP MUC21 CAPRIN1 RBL2 PHACTR4 SCARNA10 ITGB3 CCT8 HLA-C CD44 GFOD2 EPB41 GAPDH MMD C21orf7 HLA-B PTPRJ KARS PTP4A2 CHD4 SRSF1 BACH1 CLIC1 DDB1 COX4I1 KHDRBS1 RIMKLB CLTC MORC3 CFB EEF1G CYBA EIF3I A2ML1 MED13 TTC3 HLA-DRA SLC3A2 RPL13 RBBP4 ARHGDIB DDX5 BRWD1 HLA-DRB1 FAU PRPF8 SFPQ LDHB GNA13 TMPRSS2 HLA-DQB1 CFL1 SPNS2 MACF1 TWF1 PRKAR1A SIK1 HLA-DPA1 GSTP1 PFN1 YBX1 ANO6 RPL38 CSTB HLA-DPB1 CCND1 ALOX12 SLC2A1 TUBA1B EVPL PFKL RPS18 CTTN EIF5A RPS8 TMBIM6 9-Sep COL18A1 RPL10A RAB6A RPL26 PRDX1 DAZAP2 MYL12A COL6A2 SRSF3 RPS3 NCOR1 CMPK1 KRT80 MYL12B MAPK1 C6orf132 PICALM UBB RNF11 KRT4 ANKRD12 MIF HSP90AB1 BIRC3 C17orf76-AS1 NRD1 KRT78 RAB31 FBXO7 CRISP3 BIRC2 RPL23A USP24 KRT8 ROCK1 MYH9 EEF1A1 TMEM123 RPL23 JAK1 EIF4B ATP5A1 EIF3D NT5E MMP1 RPL19 PDE4B PFDN5 C18orf25 LGALS1 SYNCRIP ATM NR1D1 SERBP1 ITGA5 SMAD2 EIF3L UBE2J1 CD24 CHD1 SH3GLB1 RPL41 RAB27B DDX17 AIM1 ARPC2 PJA2 GBP2 MYL6 FECH JOSD1 SNX3 DOCK10 CDC42SE2 RPL5 PTGES3 LMAN1 RPL3 CD164 TRIP12 SKP1 SCARNA2 NACA CD226 ATF4 SERINC1 SP100 PITX1 SORT1 LRP1 HMHA1 XRCC6 VNN1 NCL TGFBI CAPZA1 CTDSP2 GPX4 ARL8B RPS12 PTMA HSPA9 RHOC USP15 MIDN EMC3 SGK1 C2orf54 MATR3 HIPK1 RAP1B RPS15 RPL32 UTRN HDLBP PAIP2 CSDE1 LYZ OAZ1 ANKRD28 SASH1 2-Sep SPINK5 ATP1A1 NAP1L1 EEF2 UBE2E1 SYNE1 RASSF2 GRPEL2 2020373103   25 Mar 2026 MAN1A2 OSBPL8 UBXN6 RPL15 SOD2 XRN2 CD74 FAM46C BTG1 RPL36 TOP2B WTAP RBM39 RPS14 TXNIP SLC25A3 RPS28 TGFBR2 IGF2R LINC00657 ANXA6 ECM1 HSP90B1 ZNF426 CMTM6 QKI TGM2 GM2A MCL1 C12orf75 CDC37 GOLGA4 RNASET2 TOP1 SPARC CDC42SE1 GLTP ILF3 RPSA MAFK SRSF6 CYFIP2 S100A10 RPL6 CALR RPL14 ACTB PKIG CLINT1 FLG PTPN11 RPL18A CTNNB1 EIF2AK1 YWHAB DOCK2 CRNN RPLP0 FKBP8 HIGD1A HNRNPA2B1 STK4 NPM1 CRCT1 RAN UBA52 GPX1 CBX3 MMP9 ATP6V0E1 LCE3E ZMYM2 UBA2 RHOA C7orf41 FAM210B HNRNPH1 LCE3D USP12 SPINT2 RPL29 7-Sep RTFDC1 CANX SPRR3 HMGB1 NCCRP1 TKT PPIA RBM38 GNB2L1 SPRR2D HSPH1 RPS19 ARF4 UPP1 RAB22A CDYL SPRR2A UFM1 ARHGEF1 ARL6IP5 CHCHD2 GNAS F13A1 SPRR2E ELF1 CNFN FOXP1 AKAP9 TUBB1 SDPR S100A8 TSC22D1 VASP CGGBP1 GNG11 RPS6KA3 STK17B S100A6 TPT1 SNRPD2 PCNP GNB2 EIF2S3 SF3B1 RPS27 LCP1 CALM3 RPL24 SERPINE1 CYBB HSPD1 TPM3 LMO7 GLTSCR2 BBX PRKAR2B DDX3X CLK1 ASH1L MYCBP2 FTL FSTL1 LAMB1 SLC9A7 ACSL1 ARHGEF2 DNAJC3 RPL13A CSTA TES TIMP1 CCT5 LMNA IPO5 RPS11 ITGB5 CAPZA2 RBM3 SUB1 CCT3 COL4A2 VSIG10L SEC61A1 SND1 MAGED2 IL7R HDGF CUL4A ZNF83 RPN1 HILPDA ALAS2 RPL37 TAGLN2 RPPH1 RPS9 CNBP CALU MSN IL6ST COPA PNP EPS8L1 CDV3 CPA4 OGT MAP3K1 UHMK1 HNRNPC RPL28 COPB2 MKLN1 RPS4X KIF2A PRRC2C ARHGAP5 RPS5 ZBTB38 CALD1 XIST TNPO1 RABGAP1L BAZ1A RPS7 XRN1 JHDM1D ATRX BTF3 TOR1AIP2 RPS29 YWHAQ PLOD2 MKRN1 SH3BGRL VCAN QSOX1 RPL36AL ODC1 TSC22D2 ATG9B MORF4L2 PTGS1 GLUL NIN PDIA6 SERP1 INSIG1 ACSL4 RPL35 LAMC1 KTN1 LPIN1 SIAH2 CTSB DOCK11 RPL12 IVNS1ABP SYNE2 OST4 MBNL1 PCM1 PGRMC1 SET TPR ACTN1 LBH SEC62 ASAH1 RPL39 AIF1L RGS18 KIAA0247 BIRC6 TBL1XR1 BNIP3L LAMP2 SETX NUCKS1 TTC9 LTBP1 AP2M1 DPYSL2 STAG2 RPL7A C1orf116 MAP3K9 EML4 PSMD2 CLU MBNL3 GPSM1 CD55 PCNX CALM2 EIF4A2 TMEM66 FLNA PLCXD1 CAPN2 TMED10 PSME4 LPP PCMTD1 RPL10 SLC25A6 ARF1 TC2N SPTBN1 ATP13A3 RPS20 MPP1 TMSB4X GUK1 PAPOLA RPS27A MUC4 SDCBP GLUD1 RPS24 TOMM20 PPP2R5C SMEK2 TFRC ASPH PTEN GHITM LYST DYNC1H1 USP34 FYTTD1 TRAM1 PCGF5 FAM25A HNRNPU EIF5 XPO1 RPL35A RPL7 TM9SF3 MORF4L1 TRIM58 THBS1 ACTR2 MRFAP1 CA2 GSTO1 BCL2A1 PFKP CHAC1 ANXA4 WDR1 UQCRB SHOC2 RPS15A NET1 SNAP23 MXD1 RPL9 RPL30 ACSL5 C2orf88 GDI2 PDIA3 ZNF638 PDS5A EIF3E ABLIM1 GLS ATP5C1 SERF2 MOB1A FRYL EIF3H RPLP2 MYO1B OPTN B2M MTHFD2 TMEM165 FAM83A CTSD RPS3A FAM107B DUOX2 TMSB10 IGFBP7 FBXO32 HBB MFAP3L RSU1 DUOXA2 TGOLN2 TMPRSS11B PSCA KIAA1199 HPGD VIM COPS2 MAL MOB1B EEF1D RHCG AKNA PIP4K2A ARPP19 RPL31 PF4 PLIN2 CIB1 ATP6V1G1 YME1L1 CGNL1 LIMS1 PPBP RPS6 IQGAP1 STOM 2020373103   25 Mar 2026 ZEB1 ANXA2 RANBP2 CCNG2 VCP HBA2 XRCC5 ITGB1 VPS13C SOWAHC HNRPDL RMRP HBA1 RPL37A HNRNPF TPM1 ACTR3 SNCA FAM108B1 RPS2 TNS1 NCOA4 RAB8B GYPC NFKB1 VPS13A PRSS22 VCL SRGN HERC1 CXCR4 UBE2D3 HNRNPK PPL PKM PPA1 PPIB ZEB2 RPL34 ISCA1 SNN NCKAP1 PSAP DENND4A SP3 PDE5A CTSL1 GSPT1 SMARCA5 P4HA1 RPL4 HNRNPA3 ANXA5 HEMGN FAM126B PTBP3 SYNPO2L RPLP1 NFE2L2 KIAA1109 RAD23B TRAK2 EEF1B2

[0151] Additional gene targets that is used as biomarkers for use with a fluid sample from the vaginal cavity of a subject are provided in Table 3. Table 3: Additional gene targets ADDITIONAL GENE TARGETS VEGFA ANGPTL6 CLEC12A ELFN2 IFNA16 LOXL1 PLXNB3 SERPINI1 VEGFB ANGPTL7 CLEC12B EMCN IFNA17 LOXL2 PLXNC1 SERPINI2 TGFB1 ANXA1 CLEC14A EPGN IFNA2 LOXL3 PLXND1 SFRP1 EGF ANXA10 CLEC17A EPO IFNA21 LOXL4 PPBP SFRP2 PTGES2 ANXA11 CLEC18A EREG IFNA4 LPA PRL SFRP4 PTGFR ANXA13 CLEC18B F10 IFNA5 LTA PROL1 SFRP5 MMP1 ANXA2 CLEC18C F12 IFNA6 LTB PRSS1 SFTA2 MMP2 ANXA3 CLEC19A F13A1 IFNA7 MASP1 PRSS12 SFTA3 MMP3 ANXA4 CLEC1A F13B IFNA8 MASP2 PRSS2 SFTPA1 ILF3 ANXA5 CLEC1B F2 IFNB1 MBL2 PRSS3 SFTPA2 MMP24 ANXA6 CLEC2A F7 IFNE MDK PSPN SFTPB MMP25 ANXA7 CLEC2B F9 IFNG MEGF10 PTN SFTPC MMP7 ANXA8 CLEC2D FAM20A IFNK MEGF11 PZP SFTPD MMP8 ANXA8L1 CLEC2L FAM20B IFNW1 MEGF6 REG1A SHH MMP9 ANXA9 CLEC3A FAM20C IGF1 MEGF8 REG1B SLPI MMP10 AREG CLEC3B FASLG IGF2 MEGF9 REG3A SPAM1 MMP11 ARTN CLEC4A FCN1 IHH MEP1A REG3G ST14 MMP12 ASTL CLEC4C FCN2 IL10 MEP1B REG4 SULF1 MMP19 BDNF CLEC4D FCN3 IL11 MMP13 RPTN SULF2 MMP26 BMP1 CLEC4E FGF1 IL12A MMP14 S100A1 TCHH HIF1A BMP10 CLEC4F FGF10 IL12B MMP15 S100A10 TCHHL1 FGF2 BMP15 CLEC4G FGF11 IL13 MMP16 S100A11 TDGF1 SERPINE1 BMP2 CLEC4M FGF12 IL15 MMP17 S100A12 TGFA PLAU BMP3 CLEC5A FGF13 IL16 MMP20 S100A13 TGFB2 COL1A1 BMP4 CLEC6A FGF14 IL17A MMP21 S100A14 TGFB3 COL2A1 BMP5 CLEC7A FGF16 IL17B MMP23B S100A16 TGM1 COL1A2 BMP6 CLEC9A FGF17 IL17C MMP27 S100A2 TGM2 COL3A1 BMP7 CNTF FGF18 IL17D MMP28 S100A3 TGM3 COL6A1 BMP8A COLEC10 FGF19 IL17F MST1 S100A4 TGM4 COL4A1 BMP8B COLEC11 FGF20 IL18 MST1L S100A5 TGM5 2020373103   25 Mar 2026 COL4A2 BRINP2 COLEC12 FGF21 IL19 MSTN S100A6 TGM6 COL17A BRINP3 CPAMD8 FGF22 IL1A MUC1 S100A7 TGM7 COL6A2 BTC CPN2 FGF23 IL1B MUC12 S100A7A THPO LAMA1 C17orf58 CRHBP FGF3 IL1F10 MUC13 S100A7L2 TIMP1 LAMB1 C1QA CRLF1 FGF4 IL1RN MUC15 S100A8 TIMP2 LAMC1 C1QB CRLF3 FGF5 IL2 MUC16 S100A9 TIMP3 LAMA4 C1QC CRNN FGF6 IL20 MUC17 S100B TIMP4 FN1 C1QL1 CSF1 FGF7 IL22 MUC19 S100G TLL1 HSPG2 C1QL2 CSF2 FGF8 IL23A MUC2 S100P TLL2 ANG C1QL3 CSF3 FGF9 IL24 MUC20 S100Z TMPRSS15 SCGB1A1 C1QL4 CSH1 FGFBP1 IL25 MUC21 SCUBE1 TNF A2M C1QTNF1 CSH2 FGFBP2 IL26 MUC22 SCUBE2 TNFSF10 A2ML1 C1QTNF2 CSHL1 FGFBP3 IL3 MUC3A SCUBE3 TNFSF11 ADAM10 C1QTNF3 CSPG4 FIGF IL34 MUC4 SDC1 TNFSF12 ADAM11 C1QTNF4 CSPG5 FLG IL36A MUC5AC SDC2 TNFSF13 ADAM12 C1QTNF5 CST1 FLG2 IL36B MUC5B SDC3 TNFSF13B ADAM15 C1QTNF6 CST11 FLT3LG IL36G MUC6 SDC4 TNFSF14 ADAM17 C1QTNF7 CST2 FREM1 IL36RN MUC7 SEMA3A TNFSF15 ADAM18 C1QTNF8 CST3 FREM2 IL37 MUC8 SEMA3B TNFSF18 ADAM19 C1QTNF9 CST4 FREM3 IL4 MUCL1 SEMA3C TNFSF4 ADAM2 C1QTNF9B CST5 FRZB IL5 NGF SEMA3D TNFSF8 ADAM20 CBLN1 CST6 FST IL6 NGLY1 SEMA3E TNFSF9 ADAM21 CBLN2 CST7 FSTL1 IL7 NODAL SEMA3F TPO ADAM22 CBLN3 CST8 FSTL3 IL9 NRG1 SEMA3G VEGFC ADAM23 CBLN4 CST9 GDF1 INHA NRG2 SEMA4A VWC2 ADAM28 CCBE1 CST9L GDF10 INHBA NRG3 SEMA4B VWC2L ADAM29 CCL1 CSTA GDF11 INHBB NRG4 SEMA4C WFIKKN1 ADAM30 CCL11 CSTB GDF15 INHBC NRTN SEMA4D WFIKKN2 ADAM32 CCL13 CSTL1 GDF2 INHBE NTF3 SEMA4F WIF1 ADAM33 CCL14 CTF1 GDF3 INS NTF4 SEMA4G WNT1 ADAM7 CCL15 CTSA GDF5 INS-IGF2 OGFOD1 SEMA5A WNT10A ADAM8 CCL16 CTSB GDF6 INSL3 OGFOD2 SEMA5B WNT10B ADAM9 CCL17 CTSC GDF7 INSL5 OSM SEMA6A WNT11 ADAMDEC1 CCL18 CTSD GDF9 INSL6 OVGP1 SEMA6B WNT16 ADAMTS1 CCL19 CTSE GDNF ISM1 P3H1 SEMA6C WNT2 ADAMTS10 CCL2 CTSF GH1 ISM2 P3H2 SEMA6D WNT2B ADAMTS12 CCL20 CTSG GH2 ITIH1 P3H3 SEMA7A WNT3 ADAMTS13 CCL21 CTSH GPC1 ITIH2 P4HA1 SERPINA1 WNT3A ADAMTS14 CCL22 CTSK GPC2 ITIH3 P4HA2 SERPINA10 WNT4 ADAMTS15 CCL23 CTSL GPC3 ITIH4 P4HA3 SERPINA11 WNT5A ADAMTS16 CCL24 CTSO GPC4 ITIH5 P4HTM SERPINA12 WNT5B ADAMTS17 CCL25 CTSS GPC5 ITIH6 PAMR1 SERPINA2 WNT6 ADAMTS18 CCL26 CTSV GPC6 ITLN1 PAPPA SERPINA3 WNT7A ADAMTS19 CCL27 CTSW GREM1 ITLN2 PAPPA2 SERPINA4 WNT7B 2020373103   25 Mar 2026 ADAMTS2 CCL28 CTSZ GRIFIN KAZALD1 PARM1 SERPINA5 WNT8A ADAMTS20 CCL3 CX3CL1 HABP2 KITLG PCSK5 SERPINA6 WNT8B ADAMTS3 CCL3L3 CXCL1 HBEGF KNG1 PCSK6 SERPINA7 WNT9A ADAMTS4 CCL4 CXCL10 HCFC1 KY PDGFA SERPINA9 WNT9B ADAMTS5 CCL4L1 CXCL11 HCFC2 LEFTY1 PDGFB SERPINB1 XCL1 ADAMTS6 CCL4L2 CXCL12 HGF LEFTY2 PDGFC SERPINB10 XCL2 ADAMTS7 CCL5 CXCL13 HGFAC LEP PDGFD SERPINB11 ZFP91 ADAMTS8 CCL7 CXCL14 HHIP LGALS1 PF4 SERPINB12 ADAMTS9 CCL8 CXCL2 HMSD LGALS12 PF4V1 SERPINB13 ADAMTSL1 CD109 CXCL3 HPSE LGALS13 PGF SERPINB2 ADAMTSL2 CD209 CXCL5 HPSE2 LGALS14 PI3 SERPINB3 ADAMTSL3 CELA1 CXCL6 HPX LGALS16 PIK3IP1 SERPINB4 ADAMTSL4 CELA2A CXCL8 HRG LGALS2 PLAT SERPINB5 ADAMTSL5 CELA2B CXCL9 HRNR LGALS3 PLG SERPINB6 AGT CELA3A DHH HTRA1 LGALS4 PLOD1 SERPINB7 AMBP CELA3B EBI3 HTRA3 LGALS7 PLOD2 SERPINB8 AMH CFC1 EDA HTRA4 LGALS8 PLOD3 SERPINB9 ANGPT1 CFC1B EGFL6 HYAL1 LGALS9 PLXDC1 SERPINC1 ANGPT2 CHRD EGFL7 HYAL2 LGALS9B PLXDC2 SERPIND1 ANGPT4 CHRDL1 EGFL8 HYAL3 LGALS9C PLXNA1 SERPINE2 ANGPTL1 CHRDL2 EGLN1 HYAL4 LGALSL PLXNA2 SERPINE3 ANGPTL2 CLC EGLN2 IFNA1 LIF PLXNA3 SERPINF1 ANGPTL3 CLCF1 EGLN3 IFNA10 LMAN1 PLXNA4 SERPINF2 ANGPTL4 CLEC10A ELANE IFNA13 LMAN1L PLXNB1 SERPING1 ANGPTL5 CLEC11A ELFN1 IFNA14 LOX PLXNB2 SERPINH1 miRNA target hsa-let-7a-5p hsa-miR- 563 hsa-miR-519a-5p hsa-miR- 4269 hsa-miR- 4663 hsa-miR-513c-3p hsa-miR-6801-3p hsa-let-7b-5p hsa-miR- 566 hsa-miR-519b-5p hsa-miR- 4277 hsa-miR-4666a-5p hsa-miR-548t-3p hsa-miR-6803-3p hsa-let-7c-5p hsa-miR- 569 hsa-miR-520c-5p hsa-miR- 4279 hsa-miR-4680-5p hsa-miR-548x-5p hsa-miR-6805-3p hsa-let-7e-5p hsa-miR- 573 hsa-miR-518d-5p hsa-miR- 4283 hsa-miR-4680-3p hsa-miR-548o-5p hsa-miR-6810-5p hsa-let-7f-5p hsa-miR-579-3p hsa-miR-1224-3p hsa-miR- 4284 hsa-miR-4685-3p hsa-miR-548aj-5p hsa-miR- 6811-5p hsa-miR-17-3p hsa-miR-584-5p hsa-miR-1225-3p hsa-miR- 4329 hsa-miR- 4686 hsa-miR-548am-5p hsa-miR-6814-5p hsa-miR-20a-5p hsa-miR- 586 hsa-miR-1228-5p hsa-miR-3605-3p hsa-miR-4687-5p hsa-miR-3529-3p hsa-miR-6817-5p hsa-miR-21 -5p hsa-miR- 600 hsa-miR-1229-3p hsa-miR-3613-5p hsa-miR-4690-5p hsa-miR-365b-3p hsa-miR-6819-5p hsa-miR-24-3p hsa-miR- 609 hsa-miR-1231 hsa-miR-3613-3p hsa-miR-4703-3p hsa-miR-1227-5p hsa-miR-6823-5p hsa-miR-26a-5p hsa-miR- 612 hsa-miR- 1264 hsa-miR-3620-3p hsa-miR-4704-3p hsa-miR-1229-5p hsa-miR-6828-5p hsa-miR-26b-5p hsa-miR- 622 hsa-miR-320b hsa-miR-3622b-3p hsa-miR- 4710 hsa-miR-1236-5p hsa-miR-6780b-5p hsa-miR-29a-3p hsa-miR- 626 hsa-miR-320c hsa-miR- 3651 hsa-miR- 4721 hsa-miR-1238-5p hsa-miR-6839-5p 2020373103   25 Mar 2026 hsa-miR-32-5P hsa-miR-628-3p hsa-miR- 1323 hsa-miR- 3656 hsa-miR-4723-5p hsa-miR-3620-5p hsa-miR-6839-3p hsa-miR-93-5P hsa-miR- 630 hsa-miR- 1301-3p hsa-miR- 3657 hsa-miR-4725-3p hsa-miR-4743-3p hsa-miR-6853-3p hsa-miR-98-5P hsa-miR-631 hsa-miR-1179 hsa-miR-3658 hsa-miR-4731-3p hsa-miR- 6068 hsa-miR-6858-5p hsa-miR-29b-3P hsa-miR-642a-5p hsa-miR-1182 hsa-miR-3663-3p hsa-miR-4735-5p hsa-miR- 6071 hsa-miR-6866-3p hsa-miR-199a-5p hsa-miR- 646 hsa-miR- 1200 hsa-miR-3670 hsa-miR-3591-3p hsa-miR- 6080 hsa-miR-6872-3p hsa-miR-181a-5p hsa-miR- 647 hsa-miR-1207-5p hsa-miR-3679-5p hsa-miR-4745-5p hsa-miR- 6083 hsa-miR-6874-5p hsa-miR-181c-5p hsa-miR- 648 hsa-miR- 548j-5p hsa-miR-3679-3p hsa-miR-4746-3p hsa-miR- 6084 hsa-miR-6876-3p hsa-miR-199b-5p hsa-miR-548d-3p hsa-miR- 1297 hsa-miR-3682-3p hsa-miR-4747-5p hsa-miR- 6089 hsa-miR-6879-5p hsa-miR-214-3P hsa-miR-653-5p hsa-miR-1244 hsa-miR- 3684 hsa-miR- 4748 hsa-miR- 6124 hsa-miR-6879-3p hsa-miR-216a-5p hsa-miR-656-3p hsa-miR- 1248 hsa-miR- 3909 hsa-miR-499b-3p hsa-miR- 6128 hsa-miR-6882-5p hsa-let-7g-5p hsa-miR-549a hsa-miR-548n hsa-miR- 3924 hsa-miR-4758-5p hsa-miR- 6132 hsa-miR-6884-5p hsa-let-7i-5p hsa-miR-660-5p hsa-miR-1268a hsa-miR-3935 hsa-miR-4760-5p hsa-miR- 6133 hsa-miR-6884-3p hsa-miR-1 hsa-miR-363-5p hsa-miR- 548i hsa-miR- 3938 hsa-miR-4762-5p hsa-miR- 6134 hsa-miR-6886-3p hsa-miR-138-5P hsa-miR-767-5p hsa-miR-1252-5p hsa-miR-374c-5p hsa-miR-4763-5p hsa-miR-548ay-5p hsa-miR-6894-5p hsa-miR-140-5P hsa-miR-769-3p hsa-miR-664a-3p hsa-miR-548aa hsa-miR-4764-3p hsa-miR-6505-5p hsa-miR-6895-5p hsa-miR-141-3P hsa-miR- 802 hsa-miR-1197 hsa-miR-1268b hsa-miR-4769-5p hsa-miR-6512-3p hsa-miR-6895-3p hsa-miR-143-3P hsa-let-7a-3P hsa-miR-320d hsa-miR-548ab hsa-miR-4769-3p hsa-miR-6515-5p hsa-miR-7113-3p hsa-miR-145-5P hsa-let-7f-1-3p hsa-miR-1908-5p hsa-miR-4419a hsa-miR-4774-3p hsa-miR-6715b-3p hsa-miR- 7114-5p hsa-miR-152-3P hsa-miR-27a-5p hsa-miR- 1910-5p hsa-miR- 4425 hsa-miR- 4775 hsa-miR-6721-5p hsa-miR-7153-3p hsa-miR-9-5p hsa-miR-92a-1-5p hsa-miR- 2113 hsa-miR- 4429 hsa-miR-4778-3p hsa-miR-6722-3p hsa-miR-7156-3p hsa-miR-206 hsa-miR-92a-2-5p hsa-miR- 759 hsa-miR- 4430 hsa-miR-4436b-3p hsa-miR-892c-5p hsa-miR-7157-3p hsa-miR-320a hsa-miR-29b-2-5p hsa-miR- 2114-5p hsa-miR- 4434 hsa-miR-4789-3p hsa-miR-134-3p hsa-miR-7159-5p hsa-miR-29c-3P hsa-miR-105-3p hsa-miR- 2115-3p hsa-miR- 4435 hsa-miR-4800-3p hsa-miR-383-3p hsa-miR-7159-3p hsa-miR-200a-3p hsa-miR- 16-2-3p hsa-miR-2276-3p hsa-miR-4436a hsa-miR- 5047 hsa-miR-597-3p hsa-miR-7160-5p hsa-miR-365a-3p hsa-miR-214-5p hsa-miR- 2278 hsa-miR- 4437 hsa-miR-5001-3p hsa-miR-605-3p hsa-miR- 7161-3p hsa-miR-302c-5p hsa-miR-23b-5p hsa-miR-711 hsa-miR-4445-5p hsa-miR-5002-5p hsa-miR-653-3p hsa-miR-7845-5p hsa-miR-369-3P hsa-miR-135a-3p hsa-miR-2681-5p hsa-miR- 4448 hsa-miR-5004-5p hsa-miR-548f-5p hsa-miR-7849-3p hsa-miR-374a-5p hsa-miR-127-5p hsa-miR- 3115 hsa-miR- 4450 hsa-miR-548ap-5p hsa-miR-1537-5p hsa-miR-7855-5p 2020373103   25 Mar 2026 hsa-miR-377-3P hsa-miR-149-3p hsa-miR-3120-3p hsa-miR- 4452 hsa-miR-548ap-3p hsa-miR-2276-5p hsa-miR- 8055 hsa-miR-379-5P hsa-miR-150-3p hsa-miR-3126-5p hsa-miR- 4456 hsa-miR-5008-5p hsa-miR-500b-3p hsa-miR- 8057 hsa-miR-330-3P hsa-miR-188-3p hsa-miR-3127-5p hsa-miR-3155b hsa-miR-5009-3p hsa-miR-6720-5p hsa-miR- 8068 hsa-miR-324-5P hsa-miR-195-3p hsa-miR- 3132 hsa-miR- 4483 hsa-miR- 5094 hsa-miR-6730-3p hsa-miR- 8070 hsa-miR-339-5P hsa-miR-194-3p hsa-miR- 466 hsa-miR- 4491 hsa-miR- 5190 hsa-miR-6736-3p hsa-miR- 8072 hsa-miR-433-3P hsa-miR-296-3p hsa-miR-544b hsa-miR- 4493 hsa-miR-5196-5p hsa-miR-6737-5p hsa-miR-8082 hsa-miR-452-5P hsa-miR-130b-5p hsa-miR-3140-3p hsa-miR- 4498 hsa-miR- 5572 hsa-miR-6737-3p hsa-miR- 8089 hsa-miR-409-3P hsa-miR-335-3p hsa-miR- 3141 hsa-miR-4419b hsa-miR-548ar-5p hsa-miR-6738-3p hsa-miR- 1199-5p hsa-miR-410-3P hsa-miR-146b-3p hsa-miR- 3143 hsa-miR- 4500 hsa-miR-548as-5p hsa-miR-6743-5p hsa-miR- 7973 hsa-miR-485-5P hsa-miR-505-5p hsa-miR-3145-3p hsa-miR- 4509 hsa-miR-5579-3p hsa-miR-6744-5p hsa-miR-491-5P hsa-miR-513a-3p hsa-miR- 3148 hsa-miR- 1587 hsa-miR-664b-3p hsa-miR-6750-5p hsa-miR-492 hsa-miR-574-5p hsa-miR-3155a hsa-miR-3120-5p hsa-miR-5580-3p hsa-miR-6752-5p hsa-miR-496 hsa-miR-548b-5p hsa-miR-3157-5p hsa-miR-3156-3p hsa-miR-5581-5p hsa-miR-6753-5p hsa-miR-515-5P hsa-miR-548a-5p hsa-miR-3158-3p hsa-miR-3173-5p hsa-miR-548at-5p hsa-miR-6755-3p hsa-miR-519e-5p hsa-miR-548c-5p hsa-miR- 3163 hsa-miR-3682-5p hsa-miR-5582-3p hsa-miR-6760-5p hsa-miR-519c-5p hsa-miR-625-3p hsa-miR- 1193 hsa-miR-3913-3p hsa-miR-5584-5p hsa-miR-6764-5p hsa-miR-520a-5p hsa-miR-548d-5p hsa-miR- 3179 hsa-miR-3925-3p hsa-miR-5584-3p hsa-miR-6766-5p hsa-miR-525-5P hsa-miR-654-3p hsa-miR-320e hsa-miR-3942-3p hsa-miR-5585-3p hsa-miR-6768-5p hsa-miR- 518f-5p hsa-miR-874-3p hsa-miR- 4296 hsa-miR- 3960 hsa-miR-5586-3p hsa-miR-6769a-3p hsa-miR-526a hsa-miR-891b hsa-miR- 4298 hsa-miR- 3972 hsa-miR-548au-5p hsa-miR-6773-5p hsa-miR-518c-3p hsa-miR-875-3p hsa-miR- 4300 hsa-miR- 3973 hsa-miR-5591-3p hsa-miR-6776-5p hsa-miR-520g-3p hsa-miR-744-5p hsa-miR- 4302 hsa-miR- 3977 hsa-miR-5682 hsa-miR-6777-3p hsa-miR-518d-3p hsa-miR-885-3p hsa-miR- 4311 hsa-miR- 4643 hsa-miR-548aw hsa-miR-6782-5p hsa-miR-520h hsa-miR-877-5p hsa-miR- 4323 hsa-miR- 4648 hsa-miR- 5703 hsa-miR-6783-5p hsa-miR-499a-5p hsa-miR- 924 hsa-miR- 4259 hsa-miR- 4651 hsa-miR-211-3p hsa-miR-6784-5p hsa-miR-504-5p hsa-miR- 944 hsa-miR- 4251 hsa-miR-4653-5p hsa-miR-514a-5p hsa-miR-6790-5p hsa-miR-455-5p hsa-miR-523-5p hsa-miR- 4326 hsa-miR-4653-3p hsa-miR-548g-5p hsa-miR-6792-5p hsa-miR-539-5p hsa-miR-518e-5p hsa-miR- 4262 hsa-miR- 4657 hsa-miR-1277-5p hsa-miR-6798-3p 2020373103   25 Mar 2026 hsa-miR-561-3P hsa-miR-522-5p hsa-miR- 4268 hsa-miR- 4658 hsa-miR-1306-5p hsa-miR-6801-5p Other properties of fluid samples

[0152] In some embodiments of method herein, other properties or contents of samples from a vaginal cavity, such as menstrual fluid, is measured. In some embodiments, the properties or contents include a small molecule, a microbiome, a pathogen, a metabolic marker or metabolome, an epigenetic marker or modification or epigenome, or a hormone.

[0153] In some embodiments, a fluid sample from the vaginal cavity of a subject, such as menstrual fluid, comprises a small molecule or chemical compound. In some embodiments, the small molecule differs in content, amount, or concentration in menstrual fluid from a subject experiencing HMB or AUB compared with a sample of menstrual fluid from a subject experiencing normal menstrual bleeding.

[0154] In some embodiments, the small molecule in such a sample is a molecule having a low molecular weight. In some embodiments, the small molecule has a molecular weight less than 1000 Da, less than 900 Da, less than 800 Da, less than 700 Da, less than 600 Da, less than 500 Da, less than 400 Da, less than 300 Da, less than 200 Da, or less than 100 Da. In some embodiments, the small molecule has a size less than 100 nm, less than 50 nm, less than 10 nm, or less than 1 nm.

[0155] In some embodiments, a small molecule is a drug, a supplement, a metabolite, or a monomer (e.g., a ribonucleotide, a deoxyribonucleotide, an amino acid, or a monosaccharide). In some embodiments, a small molecule is an organic molecule or an inorganic molecule. In some embodiments, a small molecule is able to regulate a biological process.

[0156] In some embodiments, a small molecule is a secondary metabolite. In some embodiments, a secondary metabolite is a natural metabolite or product. In some embodiments, a secondary metabolite is produced by a microorganism, such as a bacterium or fungus, such as a microorganism in a microbiome of the subject or a pathogen of the subject. In some embodiments, secondary metabolites includes alkaloids, glycosides, lipids, nonribosomal peptides (e.g., actinomycin-D), phenazines, natural phenols (e.g., flavonoids), polyketide, terpenes (e.g., steroids), and tetrapyrroles. In some embodiments, a microorganism responsible for producing such a secondary metabolite is in the sample, in the vaginal cavity of the subject, or elsewhere in or on the subject.

[0157] In some embodiments, a small molecule of a fluid sample from a vaginal cavity of a subject differs from another fluid sample from a vaginal cavity of another subject. In some 2020373103   25 Mar 2026 embodiments, a small molecule of a fluid sample of a vaginal cavity of a subject experiencing HMB or AUB differed from a small molecule of that of a subject experiencing normal menstrual bleeding. In some embodiments, a small molecule of a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB has a higher or lower concentration in a fluid sample from the vaginal cavity of the subject than in that of a subject experiencing normal menstrual bleeding. In some embodiments, a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB comprises one or more small molecules which are not present in that of a subject experiencing normal menstrual bleeding. In some embodiments, a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB lacks a small molecule which is found in that of a subject experiencing normal menstrual bleeding. In some embodiments, a fluid sample from the vaginal cavity of a subject experiencing HMB or AUB comprises a different small molecule composition, such as a different ratio of small molecules or a different set of small molecules, than that of a subject experiencing normal menstrual bleeding.

[0158] In some embodiments, a small molecule is detected using any acceptable method, including mass spectrometry (MS), gas chromatography (GC), liquid chromatography (LC), high performance liquid chromatography (HPLC), HPLC-MS, HPLC-MS / MS, GC-MS, GC-MS / MS, LC-MS, LC-MS / MS, or by an assay such as ELISA or an enzymatic assay.

[0159] In some embodiments, a microbiome detectable in a fluid sample from the vaginal cavity of a subject, such as menstrual fluid, comprises an ecological community of microorganisms, which comprises commensal, symbiotic, pathogenic microorganisms, or a combination thereof. In some embodiments, a microbiome comprises bacteria, fungi (e.g., yeast), viruses, archaea, protists, or a combination thereof. In some embodiments, a microbiome comprises members of an endometrial microbiome, uterine microbiome, or a vagino-cervical microbiome of a subject.

[0160] In some embodiments, a microbiome detectable in a fluid sample from the vaginal cavity of a subject, such as a menstrual fluid, is pathogen free or comprises one or more pathogens. In some embodiments, pathogens comprises microorganisms such as bacteria, fungi (e.g., yeast), viruses, archaea, protists, or a combination thereof. In some embodiments, a fluid sample has one type of pathogen or more than one type of pathogen. In some embodiments, a fluid sample has 2, 3, 4, 5, or more types of pathogens. In some embodiments, a pathogen found in a sample of a subject experiencing HMB or AUB is absent in a sample from a subject experiencing normal menstrual bleeding.

[0161] In some embodiments, a pathogen in a fluid sample from the vaginal cavity of a subject is indicative of HMB, AUB, a menstrual cycle disorder, or another disorder. In some 2020373103   25 Mar 2026 embodiments, a pathogen in a fluid sample from the vaginal cavity of a subject is causative of an infection, which is causative of a disease or condition. In some embodiments, such a pathogen causes HMB, AUB, a menstrual cycle disorder, or another disorder. In some embodiments, a pathogen causes a disease associated with HMB, AUB, a menstrual cycle disorder, or another disorder. In some embodiments, such a pathogen alters the symptoms of HMB, AUB, a menstrual cycle disorder or another disorder in a subject. In some embodiments, a pathogen in a fluid sample from the vaginal cavity of a subject is unrelated to HMB, AUB, a menstrual cycle disorder, or another disorder of the subject.

[0162] A pathogen is disruptive of a microbiome of a subject. In some embodiments, a pathogen reduces the quantity or variety of one or more microorganisms present in the microbiome detectable in a fluid sample from the vaginal cavity of a subject, such as menstrual fluid. In some embodiments, a pathogen causes such a reduction in microorganism population for example by out-competing one or more microorganisms of such a microbiome.

[0163] In some embodiments, a microbiome of a subject, such as an endometrial microbiome, uterine microbiome, or a vagino-cervical microbiome, which is detectable in a menstrual fluid sample, provides a potential source of novel biomarkers for detection of HMB or AUB.

[0164] In some embodiments, a microbiome of a fluid sample from a vaginal cavity of a subject differs from another fluid sample from a vaginal cavity of another subject. In some embodiments, the microbiome of a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB differs from a microbiome of that of a subject experiencing normal menstrual bleeding. In some embodiments, a microbiome of a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB has a higher or lower total amount of one or more microbes than that of a subject experiencing normal menstrual bleeding. In some embodiments, a microbiome of a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB comprises one or more microbes which are not present in that of a subject experiencing normal menstrual bleeding. In some embodiments, a microbiome of a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB lacks a microbe which is found in that of a subject experiencing normal menstrual bleeding. In some embodiments, a microbiome of a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB comprises a different composition, such as a different ratio of microbes or a different set of microbes, than that of a subject experiencing normal menstrual bleeding. In some embodiments, the difference between two or more microbiome, such as the difference between the microbiome of the a fluid sample from a vaginal cavity of a first subject and a second subject, wherein the second subject is 2020373103   25 Mar 2026 experiencing normal menstrual fluid, is referred to as beta-diversity, and is described or indexed, for example, using unweighted or weighted UniFrac distance metrics or a Bray-Curtis dissimilarity.

[0165] In some embodiments, an alteration in the microbiome of a subject compared with that of a subject experiencing normal menstrual bleeding is indicative of HMB, AUB, a menstrual cycle disorder, or another disorder. In some embodiments, one or more microbes in a microbiome is measured, quantified, or detected in a sample of menstrual fluid by an acceptable method, which comprises a sequencing technique, such as 16s rRNA sequencing, followed by analysis. In some embodiments, a property of a microbiome is measured or detected, such as one or more protein biomarkers, gene target biomarkers, expressed genes, genotype of organisms, phenotype of organisms, diversity of organisms, number of organisms, or other property is measured from a menstrual fluid sample of a subject.

[0166] In some embodiments, a fluid sample from the vaginal cavity, such as menstrual fluid, comprises one or more metabolic markers, which differs in content, amount, or concentration in a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB compared with a sample of menstrual fluid from a subject experiencing normal menstrual bleeding. Metabolic markers found in such a fluid sample comprises one or more drug metabolites, one or more endogenous metabolites, such as an amino acid, organic acid, nucleic acid, fatty acid, amine, sugar, vitamin, co-factor, pigment, antibiotic, other metabolites, or one or more exogenous metabolites, such as an environmental contaminant or xenobiotic.

[0167] In some embodiments, an amount or presence of a metabolite of a fluid sample from a vaginal cavity of a subject differs from another fluid sample from a vaginal cavity of another subject. In some embodiments, an amount or presence of a metabolite in a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB differs from an amount or presence of the same metabolite in the menstrual fluid of a subject experiencing normal menstrual bleeding. In some embodiments, an amount or presence of a metabolite in a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB is higher or lower in the menstrual fluid of a subject experiencing normal menstrual bleeding. In some embodiments, a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB comprises one or more metabolites which are not present in that of a subject experiencing normal menstrual bleeding. In some embodiments, a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB lacks a metabolite which is found in that of a subject experiencing normal menstrual bleeding. In some embodiments, the metabolic composition (e.g., metabolome) of a fluid sample from a 2020373103   25 Mar 2026 vaginal cavity of a subject experiencing HMB or AUB comprises a different composition, such as a different ratio of metabolites or a different set of metabolites than that of a subject experiencing normal menstrual bleeding.

[0168] In some embodiments, metabolites is detected using any acceptable method, including MS, GC, LC, HPLC, HPLC-MS, HPLC-MS / MS, GC-MS, GC-MS / MS, LC-MS, LC-MS / MS, or by an assay such as ELISA or an enzymatic assay. In some embodiments, a metabolomic analysis is performed to detect a plurality of metabolites in a fluid sample from a vaginal cavity of a subject.

[0169] In some embodiments, one or more epigenetic modifications to nucleic acid of a fluid sample from a vaginal cavity of a subject differs from another fluid sample from a vaginal cavity of another subject. In some embodiments, one or more epigenetic modifications to nucleic acid in a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB is altered compared with a fluid sample from a vaginal cavity of a subject from a subject experiencing normal menstrual bleeding. In some embodiments, an epigenetic modification not present in a sample from a subject experiencing normal menstrual bleeding is present in a sample from a subject experiencing HMB or AUB. In some embodiments, a modification which is present in a fluid sample from a vaginal cavity of a subject from a subject experiencing normal menstrual bleeding is not present in a sample from a subject experiencing HMB or AUB. Sometimes, a given epigenetic modification is present to a greater or lesser extent in a sample from a subject experiencing HMB or AUB than in a sample from a subject experiencing normal menstrual bleeding.

[0170] In some embodiments, epigenetic modifications to nucleic acid is measured in a fluid sample collected from a vaginal cavity of a subject such as a menstrual fluid sample. Epigenetic modifications referred to changes to a DNA strand, which are not changes to the DNA sequence. In some embodiments, epigenetic changes comprises methylation of the DNA as well as histone modification, acetylation, methylation, ubiquitylation, phosphorylation, sumoylation, ribosylation, or citrullination. In some embodiments, epigenetic modifications is of known gene targets. More than one type of epigenetic modification is present in a sample. In some embodiments, epigenetic modifications occur with other changes, such as changes in protein biomarkers, changes in gene target biomarkers, changes in gene expression, changes in nucleic acid content, changes in cell types present, and changes in flow rate of menstrual fluid.

[0171] In some embodiments, epigenetic modifications is detected using one or more acceptable methods. In some embodiments, for example when methylation has occurred, methylome 2020373103   25 Mar 2026 sequencing is performed. Methylated DNA methylation sites is uncovered for example by bisulfite conversion followed by sequencing or microarray analysis, or by genome wide methylation quantification combined with HPLC-UV, LC-MS / MS, and / or ELISA. In some embodiments, the extent of methylation of nucleic acid in a sample, such as the extent of methylation of one or more known gene targets, is determined for example by bisulfite conversion followed by q-RT-PCR or PCR and sequencing, or by performing a DNA enzyme digest based on a target sequence followed by q-RT-PCR or PCR and sequencing.

[0172] In some embodiments, one or more hormones of a fluid sample from a vaginal cavity of a subject differs from another fluid sample from a vaginal cavity of another subject. In some embodiments, one or more hormones in a fluid sample from a vaginal cavity of a subject experiencing HMB or AUB is altered compared with a sample of menstrual fluid from a subject experiencing normal menstrual bleeding. In some embodiments, a hormone not present in a fluid sample from a vaginal cavity of a subject experiencing normal menstrual bleeding is present in a sample from a subject experiencing HMB or AUB. In some embodiments, a hormone which is present in a sample from a fluid sample from a vaginal cavity of a subject experiencing normal menstrual bleeding is absent in a sample from a subject experiencing HMB or AUB. In some embodiments, a hormone is present to a greater or lesser extent (e.g., higher concentration or lower concentration) in a sample from a subject experiencing HMB or AUB than in a sample from a subject experiencing normal menstrual bleeding.

[0173] In some embodiments, a hormone in a fluid sample from a vaginal cavity of a subject is estrogen, follicle stimulating hormone, progesterone, human chorionic gonadotropin (hCG), or luteinizing hormone (LH). In some embodiments, hormone levels vary throughout the menstrual cycle in menstrual fluid or non-menstrual fluid samples of subjects

[0174] In some embodiments, a hormone is measured using any acceptable method. In some embodiments, a hormone is measured using GC-MS, GC-MS / MS, LC-MS, LC-MS / MS, HPLC, HPLC-MS, Western blotting, ELISA, a dot blot, an immunoassay, or another method. Other Tests

[0175] In some embodiments, other tests are performed in parallel with analysis of the menstrual fluid. In some embodiments, parallel testing allows for a more complete analysis of the health or disease status of a subject in some cases. In some embodiments, analysis of another test in addition to analysis of a menstrual fluid sample or cervicovaginal fluid sample is used to optimize the collection system or analysis of the fluid sample. In some embodiments, another 2020373103   25 Mar 2026 test is used to confirm or support the results of analysis on a fluid sample or the results of the analysis on a fluid sample is used to confirm or supplement the results of another test. In some embodiments, other tests comprise a blood test. In some embodiments, blood tests can include a glucose test, a measurement of the level of one or more hormones, genomic analysis, or sequencing analysis. In some embodiments, other tests comprise a test on a tissue sample. In some embodiments, tissue samples include a vaginal sample, an endometrial sample, a fallopian sample, an ovarian sample, an ovum sample, a cervical sample, a labial sample, a placenta sample, or a skin sample. In some embodiments, a test on a tissue sample comprises genomic analysis, sequencing analysis, or histological analysis. Flow Rate

[0176] In some cases of methods provided herein, menstrual blood is expelled from the body at a given flow rate. In some cases, subjects experiencing normal menstrual bleeding has a normal flow rate. During HMB or AUB, this flow rate is altered. In some embodiments, during HMB, flow rate is increased during a portion of up to the entire duration of the menstrual cycle. In some embodiments, some subjects experience a normal flow rate for a portion (e.g., a first portion) of the menstrual cycle and experience an increased flow rate during another portion (e.g., a second portion) of the menstrual cycle. In some embodiments, subjects has an increased flow rate for at least about 1 hour, at least about 2 hours, at least about 3 hours, at least about 4 hours, at least about 5 hours, at least about 6 hours, at least about 12 hours, at least about 18 hours, at least about 24 hours, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, or more.

[0177] In some methods, a flow rate of menstrual blood is measured. In some embodiments, a flow rate is measured as a normal flow rate, a high flow rate, or a low flow rate. In some embodiments, a normal flow rate is indicative of normal menstrual bleeding. In some embodiments, a high flow rate is indicative of HMB or AUB. In some embodiments, a low flow rate is indicative of AUB.

[0178] In some embodiments, a flow rate is compared with a pre-determined threshold flow rate to determine whether it is decreased or increased or normal compared with the threshold flow rate. In some embodiments, a predetermined threshold flow rate is determined from a reference sample, such as fluid collected from the vaginal cavity of a control subject, such as menstrual fluid collected from a subject experiencing normal menstrual bleeding, a subject experiencing HMB, or a subject experiencing AUB. In some embodiments, a threshold flow rate is a flow rate experienced by a subject during normal menstrual bleeding. In some embodiments, a threshold 2020373103   25 Mar 2026 flow rate is an upper threshold, such as a maximum flow rate measured during normal menstrual bleeding. In some embodiments, a threshold flow rate is determined from measuring the flow rate of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing normal menstrual bleeding.

[0179] In some embodiments, a flow rate is measured as the volume of menstrual blood collected from a subject over a time period. In some embodiments, the time period is predetermined, and is a portion of the duration of menstrual bleeding up to the entire duration of menstrual bleeding. In some embodiments, the time period is at least 5 minutes, 10 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 1.5 hours, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 18 hours, 24 hours, 48 hours, or 72 hours. In some embodiments, the time period is no more than 15 minutes, 30 minutes, 45 minutes, 1 hour, 1.5 hours, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 18 hours, 24 hours, 48 hours, or 72 hours. In some embodiments, a flow rate is measured as a maximum, median, mode, or average flow rate during a given menstrual cycle. In some embodiments, a range of flow rates is measured during a given menstrual cycle. In some embodiments, volume of menstrual blood collected during an elapsed time period is compared with a threshold value to determine normal menstrual bleeding, HMB, or AUB.

[0180] In some embodiments, a flow rate is measured by determining the amount of time required for a predetermined volume of menstrual blood to be collected from the subject. In some embodiments, a predetermined volume of blood is at least about 0.1 mL, about 0.2 mL, about 0.3 mL, about 0.4 mL, about 0.5 mL, about 0.6 mL, about 0.7 mL, about 0.8 mL, about 0.9 mL, about 1.0 mL, about 1.5 mL, about 2.0 mL, about 2.5 mL, about 3.0 mL, about 3.5 mL, about 4.0 mL, about 4.5 mL, about 5.0 mL, or more. In some embodiments, once a predetermined volume of blood has been detected, a flow rate is calculated for example by determining the ratio of the volume collected of a menstrual fluid sample to the time taken to collect the menstrual fluid sample. In some embodiments, the time elapsed during collection of a menstrual fluid sample of a predetermined volume is compared with a threshold value to determine normal menstrual bleeding, HMB, or AUB.

[0181] In some embodiments, a flow rate is measured on any day a subject is experiencing menstrual bleeding. In some embodiments, a flow rate is measured more than once during menstrual bleeding. Flow rate is measured on day 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 of menstrual bleeding. In some 2020373103   25 Mar 2026 embodiments, a flow rate is measured after 30 days of menstrual bleeding. In some embodiments, a flow rate is measured 1, 2, 3, 4, 5, or more times during a menstrual cycle.

[0182] In some embodiments, a flow rate is at least 0.01 mL per hour, at least 0.5 mL per hour, at least 0.1 mL per hour, at least 0.5 mL per hour, at least 1.0 mL per hour, at least 1.5 mL per hour, at least 2.0 mL per hour, at least 2.5 mL per hour, at least 3.0 mL per hour, at least 3.5 mL per hour, at least 4.0 mL per hour, at least 4.5 mL per hour, or at least 5.0 mL per hour. In some embodiments, a flow rate measured in a subject during one menstrual cycle is different than a flow rate measured in a subject during a different menstrual cycle. In some embodiments, a flow rate varies during a single menstrual cycle in a subject. In some embodiments, a flow rate measured at one time point in a menstrual cycle is the same or different than a flow rate measured at a different time point in the same menstrual cycle. In some embodiments, a subject experiencing HMB or AUB during a menstrual cycle experiences normal menstrual bleeding during a different part of the same menstrual cycle or during a different menstrual cycle. In some embodiments, a subject experiencing normal menstrual bleeding during a menstrual cycle experiences HMB or AUB during a different part of the same menstrual cycle or during a different menstrual cycle.

[0183] In some embodiments, an increased flow rate is a flow rate that is higher than a threshold flow rate. In some embodiments, a threshold flow rate is determined using data from subjects or other experimental data. In some embodiments, a threshold flow rate is a predicted value or a measured value. In some embodiments, a threshold flow rate is determined by measuring the flow rate of the menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing normal menstrual bleeding. In some embodiments, a threshold flow rate is determined by measuring the flow rate of the menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing HMB. In some embodiments, a threshold flow rate is determined by measuring the flow rate of the menstrual fluid of at least 1, 2, 3, 4, 5, 10, 15, 20, 30, 40, or 50 subjects experiencing AUB. In some embodiments, a threshold flow rate is determined by comparing the flow rate of subjects experiencing normal menstrual bleeding and HMB, or subjects experiencing normal menstrual bleeding and AUB. In some embodiments, a threshold flow rate is an average flow rate, a mean flow rate, a mode flow rate, a maximum flow rate, a minimum flow rate, an average flow rate plus 1, 2, or 3 standard deviations of a flow rate, or an average flow rate minus 1, 2, or 3 standard deviations of a flow rate.

[0184] In some embodiments, an increased flow rate is at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200% higher than a threshold flow rate. In some 2020373103   25 Mar 2026 embodiments, an increased flow rate is indicative of HMB or AUB. In some embodiments, a decreased flow rate is at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% lower than a threshold flow rate. In some embodiments, a decreased flow rate is a maximum rate of flow of a menstrual cycle which is lower than a threshold rate. In some embodiments, a decreased flow rate is indicative of AUB. In some embodiments, an increased or decreased flow rate indicates a menstrual cycle disorder.

[0185] In some embodiments, a flow rate of a fluid which is not menstrual fluid, such as cervicovaginal fluid or amniotic fluid, is measured using these methods. Buffer solutions

[0186] In some embodiments, fluid taken from the vaginal cavity is a complex, heterogeneous mixture of one or more cell populations and comprises one or more cell types. During HMB or AUB, the cell types found in fluid, such as menstrual blood, is altered. In some embodiments, a fluid sample from the vaginal cavity of a subject experiencing HMB or AUB comprises a different composition of cell types, more of at least one cell type, or less of at least one cell type than a subject experiencing normal menstrual bleeding. In some embodiments, prior to measuring an amount of a type of a cell in a sample, the sample is contacted with a buffer solution, e.g., by layering, pipetting, pouring, stirring, vortexing, or otherwise exposing the sample to the buffer solution.

[0187] In some embodiments, the buffer solution is a buffer which preserves one or more cell types found in a fluid sample collected from a vaginal cavity such as menstrual fluid or a buffer which holds one or more cell types found in menstrual fluid without affecting the integrity of the one or more cell types. In some embodiments, a buffer solution comprises LBgard (Biomatrica, San Diego), RNAgard (Biomatrica, San Diego), or other commercially available buffer solution. In some embodiments, a buffer solution is designed to preserve the integrity of protein, nucleic acid, lipids, metabolites, cell membranes, or whole cells.

[0188] In some embodiments, a buffer solution is at an acceptable pH. In some embodiments, a buffer solution is at a pH which is exactly 7. In some embodiments, a buffer solution is at a pH which is between 6.99 and 7.01, between 6.95 and 7.05, between 6.9 and 7.1, between 6.85 and 7.15, between 6.8 and 7.2, between 6.75 and 7.25, or between 6.7 and 7.3.

[0189] In some embodiments, a buffer solution has an acceptable osmolality. In some embodiments, a buffer solution has an osmolality of between 260 mOsm / kg and 320 mOsm / kg, between 260 mOsm / kg and 300 mOsm / kg, between 260 mOsm / kg and 280 mOsm / kg, between 280 mOsm / kg and 320 mOsm / kg, between 280 and 300 mOsm / kg, or between 300 mOsm / kg and 320 mOsm / kg. In some embodiments, a buffer solution has an osmolality of about 260 2020373103   25 Mar 2026 mOsm / kg, about 270 mOsm / kg, about 280 mOsm / kg, about 290 mOsm / kg, about 300 mOsm / kg, about 310 mOsm / kg, or about 320 mOsm / kg. In some embodiments, a buffer solution has an osmolality of more than 320 mOsm / kg or less than 260 mOsm / kg.

[0190] In some embodiments, a buffer solution has an acceptable viscosity. In some embodiments, a buffer solution has a viscosity of about 1 x 10-4 Pa-s, about 2 x 10-4 Pa-s, about 3 x 10-4 Pa-s, about 4 x 10-4 Pa-s, about 5 x 10-4 Pa-s, about 6 x 10-4 Pa-s, about 7 x 10-4 Pa-s, about 8 x 10-4 Pa-s, about 9 x 10-4 Pa-s, about 1 x 10-3 Pa-s, about 2 x 10-3 Pa-s, about 3 x 10-3 Pa-s,, about 4 x 10-3 Pa-s,, about 5 x 10-3 Pa-s,, about 6 x 10-3 Pa-s,, about 7 x 10-3 Pa-s,, about 8 x 10-3 Pa-s,, or about 9 x 10-3 Pa-s. In some embodiments, a buffer solution has a viscosity of between 1x10-4 Pa-s and 1x10-2 Pa-s, between 1x10-4 Pa-s and 5x10-3 Pa-s, between 1x10-4 Pa-s and 1x10-3 Pa-s, between 1x10-4 Pa-s and 5 x 10-4 Pa-s, between 5x10-4 Pa-s and 1x10-2 Pa-s, between 5x10-4 Pa-s and 5x10-3 Pa-s, between 5x10-4 Pa-s and 1x10-3 Pa-s, between 1x10-3 Pa-s and 1x10-2 Pa-s, between 1x10-3 Pa-s and 5x10-3 Pa-s, or between 5x10-3 Pa-s and 1x10-2 Pa-s. In some embodiments, a buffer solution has an osmolality that is approximately the same as the osmolality of water, the osmolality of blood, the osmolality of cervicovaginal fluid, or the osmolality of menstrual fluid.

[0191] In some embodiments, a buffer comprises a preservation solution. In some embodiments, a preservation solution is formulated to preserve a sample in whole or in part. In some embodiments, a preservation solution is formulated to preserve one or more cell types within a sample.

[0192] In some embodiments, the preservation solution described herein comprises at least one of: a preservation agent, a dissociation agent, or a combination thereof. In some embodiments, the preservation agent is a zwitterionic compound, an osmoprotectant, an apoptosis inhibitor, a non-reducing sugar or polyol, a disaccharide derivative, a chelating agent, a pH buffer, a phosphatase inhibitor, a protease inhibitor, or a combination thereof. In some embodiments, the dissociation agent is a mucolytic, an expectorant, a surfactant, a nuclease, a protease, or a combination thereof. In some embodiments, the preservation solution further comprises a spikein. In some embodiments, the preservation solution consists essentially of: a zwitterionic compound, an osmoprotectant, an apoptosis inhibitor, a non-reducing sugar or polyol, a chelating agent, a pH buffer, a phosphatase inhibitor, a protease inhibitor, a mucolytic, an expectorant, a surfactant, a nuclease, a protease, a spike-in, or any combination thereof. In some embodiments, the preservation solution comprises an agent for selective lysis of non-endometrial cells but not of endometrial cells in the sample. In some embodiments, the 2020373103   25 Mar 2026 preservation solution comprises an agent for selective lysis of a cell that is not an endometrial cell. In some embodiments, the agent for selective lysis is a dissociation agent. In some embodiments, the agent for selective lysis is the nuclease, the protease, or a combination thereof. In some embodiments, the preservation solution selectively lysed about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the non-endometrial cells in the sample. In some embodiments, the preservation solution selectively lyses about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the cells which are not endometrial cells in the sample. In some embodiments, the preservation solution further comprises a binding agent.

[0193] In some embodiments, the preservation solution comprises a zwitterionic compound. In some embodiments, the zwitterionic compound is a betaine or a betaine analog. In some embodiments, the zwitterionic compound is trimethylamino N-oxide (TMAO). In some embodiments, the zwitterionic compound is N-Tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid; 3-(N,N-bis[2-hydroxyethyl]amino)-2-hydroxypropanesulphonic acid; 3-(N-morpholino)propanesulfonic acid, 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; Tris(hydroxymethyl)aminomethane; piperazine-N,N’-bis(2-ethanesulfonic acid); 2-(N-Morpholino)ethanesulfonic acid hydrate; N,N-Bis(2-hydroxyethyl)-2-aminoethanesulfonic acid; N-[Tris(hydroxymethyl)methyl]glycine; 3-((3-acrylamidopropyl)-dimethylammonio)-propane-1-sulfonate; hydroxyectoine; ectoine; homoectoine; L-carnitine; sarcosine; N,N-Dimethylglycine triethylammonium acetate; glycerol phosphate; tricine; pentaerythritol; N-ethyl-N,N-bis-(2-hydroxyethyl)ammonium-N-4-butyl sulfonate; 3-morpholino-2-hydroxypropanesulfonic acid; 4-(2-ethoxy-2-oxoethyl)-4-ethylmorpholin-4-ium bromide; N-(2-ethoxy-2-oxoethyl)-3-hydroxy-N,N-bis(2-hydroxyethyl)propan-1-aminium bromide; 2-ethoxy-N,N,N-triethyl-2-oxoethanaminium bromide; 2-((3-hydroxypropyl)dimethylammonio)acetate; 2-((2-hydroxypropyl)dimethylammonio) acetate; 2-(2-(hydroxymethyl)-1-methylpiperidinium-1-yl)acetate; 2-((2-hydroxyethyl)dimethylammonio)acetate; 2-((2,3-dihydroxypropyl) dimethylammonio)acetate; 1-(2-ethoxy-2-oxoethyl)-4-hydroxy-1-methylpiperidinium bromide; 2-(4-hydroxy-1-methylpiperidinium-1-yl)acetate; 2-ethoxy-N-(2-(2-hydroxyethoxy)ethyl)-N,N-dimethyl-2-oxoethanaminium bromide; 2-((2-(2-hydroxyethoxy)ethyl)dimethylammonio)acetate; 2-(bis(2-hydroxyethyl)-(methyl)ammonio)acetate; 4-(2-hydroxyethyl)-4-methyl-2-oxomorpholin-4-ium bromide; 2-(bis(2-hydroxyethyl)-(methyl)ammonio)acetate; 2-(4-(2-hydroxyethyl)morpholino-4-ium)acetate; 4-(2-ethoxy-2-oxoethyl)-4-methylmorpholin-4-ium bromide; 1-(2-ethoxy-2-oxoethyl)-1-methylpyrrolidinium bromide; 2-(benzyl(2-hydroxy- 2020373103   25 Mar 2026 ethyl)(methyl)ammonio)acetate; 3-(2,3-dihydroxypropyl)-1-methyl-1H-imidazol-3-ium chloride; 1,3-dimethyl-1H-imidazol-3-ium methyl sulfate; N-benzyl-2-ethoxy-N,N-dimethyl-2-oxoethanaminium bromide; 1-(2-ethoxy-2-oxoethyl)-1-methylpiperidi-nium bromide; N-(2-ethoxy-2-oxoethyl)-N,N-dimethylbenzenaminium bromide; 1-(2-ethoxy-2-oxoethyl)-3 -hydroxy-1-methylpiperidinium bromide; 3-(2-(2-hydroxyethoxy)ethyl)-1-methyl-1H imidazol-3-ium chloride; 3-(2-(2-(2-hydroxyethoxy)ethoxy)ethyl)-1-methyl-1H-imidazol-3-ium chloride; 1-methyl-3-tetradecyl-1H-imidazol-3-ium bromide; N-(2-ethoxy-2-oxoethyl)-N,N-dimethylcyclo-hexanaminium bromide; 3-((2-hydroxy-ethyl)dimethyl-ammonio)pro-panoate; or any combination thereof. In some embodiments, the zwitterionic compound is a polyzwitterion. In some embodiments, the polyzwitterion is carboxybetaine methacrylate-1; carboxybetaine methacrylate-1-tertiary amine; carboxybetaine methacrylate-2; carboxybetaine acrylamide-2; carboxybetaine acrylamide-2-ethyl ester; carboxybetaine acrylamide-2-RGD; carboxybetaine diacrylamide crosslinker; glycine betaine; poly-sulfobetaine; or any combination thereof.

[0194] In some embodiments, the preservation solution comprises an osmoprotectant. In some embodiments, an osmoprotectant is for example trimethylammonium acetate; glycerol phosphate; diglycerol phosphate, N-(2-hydroxy-1,1-bis(hydroxymethyl)ethyl)glycine; 3-(N-morpholino)-2-hydroxypropanesulfonic acid; pentaerythritol; glyceric acid; malic acid; tartaric acid; lactic acid; glycolic acid; 2-hydroxybutyric acid; 3-hydroxybutyric acid; 4-amino-3-hydroxybutyric acid; 3-(1-azoniabicyclo[2.2.2]oct-1-yl)propane-1-sulfonate; 1-(2-carboxylatoethyl)-1-azabicyclo[2.2.2]octan-1-ium; or any combination thereof.

[0195] In some embodiments, the preservation solution comprises an apoptosis inhibitor. In some embodiments, the apoptosis inhibitor is PERK-eIF2-a inhibitor, ASK1 inhibitor, NRF2-KEAP1 inhibitor, JNK inhibitor, p38 MAP kinase inhibitor, IRE1 inhibitor, GSK3 inhibitor, PIK3 pathway inhibitor, MEK inhibitor, calpain inhibitor, caspase-1 inhibitor, or any combination thereof.

[0196] In some embodiments, the preservation solution comprises a non-reducing sugar or polyol. In some embodiments, the non-reducing sugar or polyol is glycol, glycerol, erythritol, threitol, arabitol, xylitol, ribitol, adonitol, mannitol, sorbitol, galactitol, fucitol, iditol, inositol, adonitol, sucralfate, sucrose octasulfate, sucrose, trehalose, or any combination thereof. In some embodiments, the preservation solution comprises a disaccharide derivative. In some embodiments, the disaccharide derivative comprises sucralose, trichloronated maltose, or a combination thereof. 2020373103   25 Mar 2026

[0197] In some embodiments, the preservation solution comprises a chelating agent. In some embodiments, the chelating agent is diethylenetriaminepentaacetic acid (DTPA); ethylenediaminetetraacetic acid (EDTA); ethylene glycol tetraacetic acid (EGTA); trans-1,2-diaminocyclohexane-N,N,N’,N’-tetraacetic acid (CDTA); 1,2-bis(2-aminophenoxy)ethane-N,N,N’,N’-tetraacetic acid (BAPTA); 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA); N-(2-hydroxyethyl)ethylenediamine-N,N’,N’-triacetic acid; sodium gluconate; nitrilotriacetic acid (NTA); or a combination thereof.

[0198] In some embodiments, the preservation solution comprises a pH buffer. In some embodiments, the pH buffer comprises citric acid; tartaric acid; malic acid; sulfosalicylic acid; sulfoisophthalic acid; oxalic acid; borate; CAPS (3-(cyclohexylamino)-1-propanesulfonic acid); CAPSO (3-(cyclohexylamino)-2-hydroxy-1-propanesulfonic acid); EPPS (4-(2-hydroxyethyl)-1-piperazinepropanesulfonic acid); HEPES (4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid); MES (2-(N-morpholino)ethanesulfonic acid); MOPS (3-(N-morpholino)propanesulfonic acid); MOPSO (3-morpholino-2-hydroxypropanesulfonic acid); PIPES (1,4-piperazinediethanesulfonic acid); TAPS (N[tris(hydroxymethyl)methyl]-3-aminopropanesulfonic acid); TAPSO (2-hydroxy-3-[tris(hydroxymethyl)methylamino]-1-propanesulfonic acid); TES (N-[tris(hydroxymethyl)methyl]-2-aminoethanesulfonic acid); bicine (N,N-bis(2-hydroxyethyl)glycine); tricine (N-[tris(hydroxymethyl)methyl]glycine); tris (tris(hydroxymethyl)aminomethane); bis-tris (2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)-1,3-propanediol); or a combination thereof.

[0199] In some embodiments, the preservation agent comprises a phosphatase inhibitor. In some embodiments, the phosphatase inhibitor comprises beta-Glycerophosphate, aprotinin, bestatin, EDTA, leupeptin, pepstatin A, or a combination thereof.

[0200] In some embodiments, the preservation agent comprises a protease inhibitor. In some embodiments, the protease inhibitor is (2R)-2-Mercaptomethyl-4-methylpentanoyl-beta-(2-naphthyl)-Ala-Ala Amide; 2-Antiplasmin; 3,4-Dichloroisocoumarin; 4-(2-Aminoethyl) benzenesulfonyl fluoride hydrochloride; 5-(R,S)-T-trans-Cinnamido-7-methyl-4-oxo-octanoyl-L-prolyl-L-proline ; a1-Antchymotrypsin; a1-Antitrypsin; a2-Antiplasmin; a2-Macroglobulin; Antithrombin III; Aprotinin; Bromoenol lactone; BTEE; C1 Esterase inhibitor; Chymostatin; Complement C1 esterase inhibitor; Dichloromethylenediphosphonic acid disodium salt; Diisopropyl fluorophosphate; e-Amino-n-caproic acid; Ecotin; EDTA; Eglin C fragment 60-63 methyl ester; Gabexate mesylate; Histatin 5; Ile-Pro-Ile; Isoamylphosphonyl-Gly-L-Pro-L-Ala; Leupeptin; N a-p-Tosyl-L-lysine chloromethyl ketone hydrochloride; N-Acetyl-eglin C; N- 2020373103   25 Mar 2026 Tosyl-L-phenylalanine chloromethyl ketone; p-Chloromercuribenzoic acid Free Acid; Phenylmethylsulfonyl fluoride; Trypsin Inhibitor; Trypsin-chymotrypsin inhibitor; Z-L-Phe chloromethyl ketone; Boc-Asp(OMe)-fluoromethyl ketone; Z-Ala-Glu(OMe)-Val-Asp(OMe)-fluoromethyl ketone; Antipain dihydrochloride from microbial source protease inhibitor; CA-074 methyl ester; Calpain Inhibitor I; Calpain Inhibitor II; Cystatin; E-64 protease inhibitor; Leupeptin trifluoroacetate salt; a2-Macroglobulin; Procathepsin B; Z-Leu-Leu-Leu-fluoromethyl ketone; Z-Phe-Phe-fluoromethyl ketone; or a combination thereof.

[0201] In some embodiments, the preservation solution comprises a spike-in. In some embodiments, as used herein, a “spike-in” is a molecule, such as a nucleic acid, a cell, or a set of molecules or cells added to a sample, wherein the spike-in is used to quantitatively or qualitatively assess or to normalize a sample. In some embodiments, the spike-in is a nucleic acid spike-in. In some embodiments, the nucleic acid spike-in is a DNA spike-in, an RNA spikein, a bacterial spike-in, or a combination thereof. In some embodiments, the DNA spike-in is a synthetic DNA or a plurality of synthetic DNAs. In some embodiments, the RNA spike-in is a synthetic RNA or a plurality of synthetic RNAs. In some embodiments, the RNA spike-in is a set of RNA transcripts developed by the External RNA Controls Consortium (ERCC).

[0202] In some embodiments, the preservation solution comprises a mucolytic agent. In some embodiments, the mucolytic agent dissociates (e.g., “unclump”) at least a portion of cellular aggregations in the cervicovaginal sample. In some embodiments, the mucolytic is acetylcysteine, ambroxol, bromhexine, carbocisteine, domiodol, dornase alfa, eprazinone, erdosteine, letosteine, mannitol, mesna, neltenexine, sobrerol, stepronin, tiopronin, N-acetyl-L-cysteine, L-acetyl cysteine / Liberase™, or a combination thereof.

[0203] In some embodiments, the preservation solution comprises an expectorant. In some embodiments, the expectorant is althea root, antimony pentasulfide, creosote, guaiacolsulfonate, guaifenesin (+ oxomemazine), ipecacuanha, levoverbenone, potassium iodide, senega, tyloxapol, ammonium chloride, or a combination thereof.

[0204] In some embodiments, the preservation solution comprises a surfactant. In some embodiments, the surfactant is polyoxyethylene glycol octylphenol ethers; polyoxyethylene glycol alkylphenol ethers; polyoxyethylene glycol sorbitan alkyl esters; sorbitan alkyl esters; polyethylene glycol; polypropylene glycol; carboxylates; sulphonates; petroleum sulphonates; alkylbenzenesulphonates; naphthalenesulphonates; olefin sulphonates; alkyl sulphates; sulphates; sulphated esters; sulphated alkanolamides; alkylphenols; ethoxylated aliphatic alcohol; polyoxyethylene surfactants; carboxylic esters; polyethylene glycol esters; 2020373103   25 Mar 2026 anhydrosorbitol esters; glycol esters; carboxylic amide; monoalkanolamine condensates; polyoxyethylene fatty acid amides; quaternary ammonium salts; polyoxyethylene alkyl and alicyclic amines; N,N,N’,N’ tetrakis substituted ethylenediamines; 2-alkyl 1-hydroxethyl 2-imidazolines; or a combination thereof.

[0205] In some embodiments, the preservation solution comprises a nuclease. In some embodiments, the nuclease is a Benzonase®, DNAse I, DNAse II, Exonuclease III, Micrococcal Nuclease, Nuclease P1, Nuclease S1, Phosphodiesterase I, Phosphodiesterase II, RNAse A, RNAse H, RNAse T1, or a combination thereof.

[0206] In some embodiments, the preservation solution comprises a protease. In some embodiments, the protease is adispase II, trypsin, pronase, collagenase 1, collagenase 2, collagenase 3, collagenase 4, hyaluronidase, pepsin, papain, chemotrypsin, chymase, clostripain, complement C1r, complement C1s, complement factor D, complement factor I, cucumisin, dipeptidyl peptidase, elastase, endoproteinase, enterokinase, Factor X Activated, caspase, cathepsin, matrix metalloprotease, or a combination thereof.

[0207] In some embodiments, the osmolality of the preservation solution is from about 310 to about 410 mOsm kg-1. In some embodiments, the osmolality of the preservation solution is from about 95 to about 210 mOsm kg-1.

[0208] In some embodiments, the preservation solution does not comprise a fixative. In some embodiments, the fixative comprises an alcohol, an aldehyde, an oxidizing agent, a metallic fixative or a combination thereof. In some embodiments, the alcohol is methanol, ethanol, propanol, isopropanol, butanol, or a combination thereof. In some embodiments, the aldehyde is formaldehyde, glutaraldehyde, or a combination thereof. In some embodiments, the oxidizing agent is an osmium tetraoxide, potassium permanganate, potassium dichromate, or a combination thereof. In some embodiments, the metallic fixative is a mercuric chloride, a picric acid, or a combination thereof. In some embodiments, the preservation solution does not comprise an alcohol, an aldehyde, an oxidizing agent, a metallic fixative, or a combination thereof.

[0209] In some embodiments, the preservation solution comprises a binding agent. In some embodiments, the binding agent selectively binds to an endometrial cell, a non-endometrial cell of the individual, spermatozoa, bacterial cell, fungal cell, or a combination thereof. In some embodiments, the binding agent selectively binds to at least one protein or fragment thereof. In some embodiments, the at least one protein or fragment thereof is a biomarker of endometriosis. In some embodiments, the binding agent selectively binds to nucleic acid. In some 2020373103   25 Mar 2026 embodiments, the nucleic acid is a biomarker of endometriosis. In some embodiments, the binding agent is immobilized, for example, to a bead or to a surface of a component of the systems described herein. In some embodiments, the binding agent is coupled to the bead or the surface of the system. In some embodiments, the binding agent is reversibly or irreversibly coupled to the bead or the surface of the system. In some embodiments, the binding agent comprises a cleavable moiety, for example, a cleavable linker. In some embodiments, the cleavable linker is cleaved photolytically, chemically, thermally, or enzymatically.

[0210] In certain embodiments, for example, about 0.5 ml of preservation solution (Biomatrica® LBgard™) is diluted in about 8.5 ml of distilled water to form a diluted preservation solution. In other embodiments, for example, for example, about a volume of preservation solution (Biomatrica® RNAgard™) is diluted in a volume of distilled water to form a diluted preservation solution. In some embodiments, such a diluted preservation solution is used in the methods and / or systems provided herein. In some embodiments, the diluted preservation solution is added to a tampon at about 3 ml to about 5 ml of diluted preservation solution per gram of fluid that is absorbed into the tampon. In some embodiments, a light absorbency tampon absorbs up to 6 g of fluid, thus, about 18 ml to about 30 ml of diluted preservation solution is added to the light absorbency tampon. In some embodiments, the diluted preservation solution is added to the tampon in the system described herein, following the rupture of the disruptable member. In some embodiments, accordingly, as the absorbency of the tampon increases, the amount of diluted preservation solution to be added increases. In some embodiments, the sample is incubated in the buffer solution prior to analysis. In some embodiments a sample is incubated in a buffer solution at about 4 C, about 10 C, about 15 C, about 20 C-, about 25 °C, about 30 °C, about 35 C, about 40 C, about 45 C, about 50 C, about 55 C, about 60 C, about 55 °C, about 60 C, about 65 C, about 70 C, about 75 C, about 80 C, about 85 °C, about 90 °C, about 95 °C, or about 100 C. In some embodiments, a sample is incubated in a buffer solution at between 4 °C and 100 C, between 4 C and 50 °C, between 4 C and 30 C, 4 C and 20 C, between 4 C and 15 C, between 4 C and 10 C, between 10 C and 20 °C, between 10 C and 15 °C, between 15 °C and 20 °C, between 20 C and 100 C, between 20 C and 50 °C, between 20 °C and 40 C , between 20 °C and 35 C , between 20 C and 30 C, between 20 C and 25 °C, between 25 C and 40 °C, between 25 °C and 35 C, between 25 C and 30 C, between 30 C and 40 C, between 30 C and 35 °C, between 35 °C and 40 °C, between 30 C and 100 C , between 30 C and 90 C, between 30 C and 80 C, between 30 C and 70 C, between 30 C and 60 °C, between 30 °C and 50 °C, between 40 °C and 100 C, between 40 C and 90 C, between 40 °C and 80 C, between 40 °C and 70 °C, between 40 °C 2020373103   25 Mar 2026 and 60 C, between 40 C and 50 C, between 50 C and 70 °C, between 50 °C and 60 C, between 60 C and 100 C, between 60 C and 90 C, between 60 C and 80 C, between 60 °C and 70 °C, between 70 C and 100 C, between 70 C and 90 °C, between 70 °C and 80 C, between 80 °C and 100 C, between 80 C and 90 C, or between 90 °C and 100 C.

[0211] In some embodiments, incubation of a sample lasts for at least about 30 seconds, about 1 minute, about 2 minutes, about 5 minutes, about 10 minutes, about 15 minutes, about 30 minutes, about 45 minutes, about 1 hour, about 2 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 7 hours, about 8 hours, about 9 hours, about 10 hours, about 11 hours, about 12 hours, overnight, for about 24 hours, or more. In some embodiments, incubation of a sample lasts for between 30 seconds and 24 hours, between 30 seconds and 12 hours, between 30 seconds and 6 hours, between 30 seconds and 1 hour, between 30 seconds and 30 minutes, between 30 seconds and 1 minute, between 1 minute and 1 hour, between 1 minute and 45 minutes, between 1 minutes and 30 minutes, between 1 minute and 10 minutes, between 1 minute and 5 minutes, between 5 minutes and 1 hour, between 5 minutes and 45 minutes, between 5 minutes and 30 minutes, between 5 minutes and 15 minutes, between 5 minutes and 10 minutes, between 10 minutes and 1 hour, between 10 minutes and 45 minutes, between 10 minutes and 30 minutes, between 10 minutes and 15 minutes, between 15 minutes and 1 hour, between 15 minutes and 45 minutes, between 15 minutes and 30 minutes, between 30 minutes and 24 hours, between 30 minutes and 12 hours, between 30 minutes and 6 hours, between 30 minutes and 1 hour, between 30 minutes and 45 minutes, between 45 minutes and 1 hour, between 1 hour and 24 hours, between 1 hour and 12 hours, between 1 hour and 6 hours, between 1 hour and 5 hours, between 1 hour and 4 hours, between 1 hour and 3 hours, between 1 hour and 2 hours, between 2 hours and 6 hours, between 2 hours and 5 hours, between 2 hours and 4 hours, between 2 hours and 3 hours, between 3 hours and 6 hours, between 3 hours and 5 hours, between 3 hours and 4 hours, between 4 hours and 6 hours, between 4 hours and 5 hours, between 5 hours and 6 hours, between 6 hours and 24 hours, between 6 hours and 12 hours, or between 12 hours and 24 hours.

[0212] In some embodiments, the sample is incubated in the buffer solution prior to analysis. In some embodiments, the buffer solution is incubated at a given temperature for at least 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 1 week, 2 weeks, 4 weeks, or more. In some embodiments, the sample is stored at room temperature, in a refrigerator, in a freezer, in a -80 °C freezer, on dry ice, or in liquid nitrogen. 2020373103   25 Mar 2026

[0213] In some embodiments, the sample is contacted with the buffer solution at a ratio of about 10% v / v, 20% v / v, 30% v / v, 40% v / v, 50% v / v, 60% v / v, 70% v / v, 80% v / v, 90% v / v, 100% v / v, 150% v / v, 200% v / v, 250% v / v, 300% v / v, 400% v / v, 500% v / v, 600% v / v, 700 % v / v, 800% v / v, 900% v / v, or 1000% v / v, where v / v indicates the ratio of the volume of the buffer solution to the volume of the sample). In some embodiments, a sample is contacted with the buffer solution at a ratio of less than 10% v / v or greater than 1000% v / v. In some embodiments, the sample is contacted with the buffer solution at a ratio of between 10% v / v and 1000% v / v, between 10% v / v and 900% v / v, between 10% v / v and 800% v / v, between 10% v / v and 700% v / v, between 10% v / v and 600% v / v, between 10% v / v and 500% v / v, between 10% v / v and 400% v / v, between 10% v / v and 300% v / v, between 10% v / v and200% v / v, between 10% v / v and 100% v / v, between 100% v / v and 1000% v / v, between 100% v / v and 900% v / v, between 100% v / v and 800% v / v, between 100% v / v and 700% v / v, between 100% v / v and 600% v / v, between 100% v / v and 500% v / v, between 100% v / v and 400% v / v, between 100% v / v and 300% v / v, between 100% v / v and 200% v / v, between 200% v / v and 100% v / v, between 200% v / v and 900% v / v, between 200% v / v and 800% v / v, between 200% v / v and 700% v / v, between 200% v / v and 600% v / v, between 200% v / v and 500% v / v, between 200% v / v and 400% v / v, between 200% v / v and 300% v / v, between 300% v / v and 1000% v / v, between 300% v / v and 900% v / v, between 300% v / v and 800% v / v, between 300% v / v and 700% v / v, between 300% v / v and 600% v / v, between 300% v / v and 500% v / v, between 300% v / v and 400% v / v, between 400% v / v and 1000% v / v, between 400% v / v and 900% v / v, between 400% v / v and 800% v / v, between 400% v / v and 700% v / v, between 400% v / v and 600% v / v, between 400% v / v and 500% v / v, between 500% v / v and 1000% v / v, between 500% v / v and 900% v / v, between 500% v / v and 800% v / v, between 500% v / v and 700% v / v, between 500% v / v and 600% v / v, between 600% v / v and 1000% v / v, between 600% v / v and 900% v / v, between 600% v / v and 800% v / v, between 600% v / v and 700% v / v, between 700% v / v, and 1000% v / v, between 700% v / v and 900% v / v, between 700% v / v and 800% v / v, between 800% v / v and 1000% v / v, between 800% v / v and 900% v / v, or between 900% v / v and 1000% v / v. In some embodiments, the sample is contacted with the buffer solution at a ratio of between 10% v / v and 100% v / v, between 10% v / v and 90% v / v, between 10% v / v and 80% v / v, between 10% v / v and 70% v / v, between 10% v / v and 60% v / v, between 10% v / v and 50% v / v, between 10% v / v and 40% v / v, between 10% v / v and 30% v / v, between 10% v / v and 20% v / v, between 20% v / v and 100% v / v, between 20% v / v and 90% v / v, between 20% v / v and 80% v / v, between 20% v / v and 70% v / v, between 20% v / v and 60% v / v, between 20% v / v and 50% v / v, between 20% v / v and 40% v / v, between 20% v / v 2020373103   25 Mar 2026 and 30% v / v, between 30% v / v and 100% v / v, between 30% v / v and 90% v / v, between 30% v / v and 80% v / v, between 30% v / v and 70% v / v, between 30% v / v and 60% v / v, between 30% v / v and 50% v / v, between 30% v / v and 40% v / v, between 40% v / v and 100% v / v, between 40% v / v and 90% v / v, between 40% v / v and 80% v / v, between 40% v / v and 70% v / v, between 40% v / v and 60% v / v, between 40% v / v and 50% v / v, between 50% v / v and 100% v / v, between 50% v / v and 90% v / v, between 50% v / v and 80% v / v, between 50% v / v and 70% v / v, between 50% v / v and 60% v / v, between 60% v / v and 100% v / v, between 60% v / v and 90% v / v, between 60% v / v and 80% v / v, between 60% v / v and 70% v / v, between 70% v / v and 100% v / v, between 70% v / v and 90 % v / v, between 70% v / v and 80 % v / v, between 80% v / v, and 100% v / v, between 80 % v / v and 90 % v / v, or between 90 % v / v and 100 % v / v.

[0214] In some embodiments, upon contact with the buffer, cell membranes of at least about 80%, 85%, 90%, 95%, 99%, or 100% of cells present in the sample remains in a substantially intact state. In some embodiments, an intact state in some cases comprises an absence of pores or tears in the cell membrane, an absence of disruption of glycosylation (e.g., glycosylation of surface molecules), or an absence of denaturing (e.g. denaturation of surface proteins).

[0215] In some embodiments, upon contact with the buffer, the cell membranes of at least 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of the types of cells in the sample is maintained in a substantially intact state. In some embodiments, a cell membrane of a type of cell is considered to be maintained in a substantially intact state if at least about 80%, 85%, 90%, 95%, 99%, or 100% of cells of that type present in the sample remain in a substantially intact state. Device for collection or measurement of menstrual fluid

[0216] In some embodiments of methods provided herein, a sample collection device is used for collection of a menstrual fluid sample from a subject. In some embodiments, any device capable of collecting a sample from a subject is used. In some embodiments, a sample collection device comprises means of collecting menstrual fluid. In some embodiments, a sample collection device additionally comprises other components to measure, store, preserve, or ship a sample. In some embodiments, a sample collection device is provided with one or more lab accessories, such as a syringe or a vacutainer, to allow for removal of a sample from the device or preparation or analysis of the sample.

[0217] In some embodiments, a sample collection device hold a volume of menstrual fluid. In some embodiments, a sample collection device has a capacity of at least 0.5 mL, 1.0 mL, 1.5 mL, 2.0 mL, 2.5 mL, 3.0 mL, 3.5 mL, 4.0 mL, 4.5 mL, or 5.0 mL. In some embodiments, a volume of fluid collected in a sample collection device is equal to or less than the capacity of the 2020373103   25 Mar 2026 sample collection device. In some embodiments, up to 0.5 mL, 1.0 mL, 1.5 mL, 2.0 mL, 2.5 mL, 3.0 mL, 3.5 mL, 4.0 mL, 4.5 mL, or 5.0 mL of menstrual fluid is collected in a sample collection device. In some embodiments, at least 0.5 mL, 1.0 mL, 1.5 mL, 2.0 mL, 2.5 mL, 3.0 mL, 3.5 mL, 4.0 mL, 4.5 mL, or 5.0 mL of menstrual fluid is collected in a sample collection device.

[0218] In some embodiments, a sample collection device comprises a means of collecting menstrual fluid from a subject, such as a tampon. In some embodiments, a tampon is inserted into the vagina of a subject, and menstrual fluid collects in or on the tampon. In some embodiments, after insertion into the vagina of a subject, time is allowed to elapse such that a sufficient volume of menstrual fluid is collected. In some embodiments, after an amount of time has passed, the tampon is removed.

[0219] In some embodiments, another means of collecting menstrual fluid, either inside the vagina or external to the vagina, is used, such as a pad, a tampon, a vaginal cup, a cervical cap, a menstrual disk, a cervical disk, a sponge, or an interlabial pad. In some embodiments, these, or other means, of collecting menstrual fluid is interchangeable.

[0220] In some embodiments, a sample collection device further comprises a collection packet. In some embodiments, a collection packet is a device used to receive a tampon or other means of collecting menstrual fluid from a subject. In some embodiments, a collection packet is a cylindrical device or is any shape which accommodates a tampon or other means of collecting menstrual fluid inside.

[0221] In some embodiments, a sample collection packet has a capacity of at least the volume of the means of collecting menstrual fluid. In some embodiments, a sample collection packet has a capacity of at least 0.5 mL, 1.0 mL, 1.5 mL, 2.0 mL, 2.5 mL, 3.0 mL, 3.5 mL, 4.0 mL, 4.5 mL, 5.0 mL, 6.0 mL, 7.0 mL, 8.0 mL, 9.0 mL, 10.0 mL, or more.

[0222] In some embodiments, a sample collection packet comprises a lid. In some embodiments, a lid is snapped closed, screwed closed, closed using a vacuum force, taped closed, glued closed, or crimped closed. In some embodiments, a lid is removable or non-removable once it is closed. In some embodiments, a sample collection packet is closed without a lid, for example by pinching the top of the sample collection packet or by using other closing means such as a zipper mechanism or an adhesive.

[0223] In some embodiments, a sample collection device comprise a device or kit as described in WO 2016 / 025332 A1 or WO 2017 / 180909 A1. In some embodiments, a representative and exemplary system includes at least some of the following components: a specialized sample collector which optimizes collection of fluid from a vaginal cavity (e.g., menstrual fluid) for 2020373103   25 Mar 2026 testing; a biological matrix extractor comprising a compression top which pull fluid, vaginal mucosa, or semen into an assay delivery reservoir, through a filter; an assay cartridge which evaluates the biological content of a biological matrix; a cartridge reader which automates assay development, result capture, and / or result interpretation; or a mobile app interface that interprets and / or track a user’s results and curates validated recommendations for health and behavior. In some embodiments, the cartridge reader electronically interfaced with a mobile phone by plugging into a headphone jack on the mobile phone. Optionally, a web-based interface provides access to easy interventions like food shopping, vitamin stores, and health facilities for therapeutics, and provides a positive behavioral feedback loop to increase prevention adherence.

[0224] In some embodiments, a sample collection device comprises a system for collecting a biological sample from a subject, which comprises a comprising a sample collector that non-invasively collects the biological sample from the subject. In some embodiments, the sample collector is inserted into ta subject’s vaginal cavity to collect the biological sample. In some embodiments, the system described herein collects a volume of biological sample comprising menstrual fluid, cervicovaginal fluid, secreted mucus, shed uterus cells, shed ovary cells, or other cells, tissue, or fluid. In some embodiments, the sample collector is made of materials that are capable of collecting and / or retaining the biological sample. In some embodiments, the sample collector is made of highly absorbent materials that absorb a liquid sample rapidly. In some embodiments, the sample collector is made of materials that release absorbed liquid samples rapidly, such as when a compression mechanism (e.g., pressure, force) is applied to the sample collector. In various embodiments, disposing the menstrual fluid sample in a preservation solution to form the mixture comprises placing a sample collector into a first central cavity of a system wherein the sample collector is compressed or squeezed, for example, to remove at least a portion of the sample from the sample collector. In some embodiments, the system comprises an extractor for extracting the biological sample from the sample collector. In some embodiments, the extractor comprises a component for applying a compression mechanism to the sample collector. In some embodiments, components for applying compression mechanisms include but are not limited to a spring, threaded screw, lever, air-tight plunger, or roller-based compression. In some embodiments, the liquid sample absorbed on a sample collector is extracted by applying a compression mechanism to the sample collector. In some embodiments, the system comprises the compression mechanism. In some embodiments, the system does not comprise a compression mechanism. In some embodiments, the compression mechanism is compressed outside of the system. In some embodiments, closing or 2020373103   25 Mar 2026 sealing the system activates the compression mechanism. In some embodiments, closing or sealing the system does not activate the compression mechanism. In some embodiments, the compression mechanism is activated separately from closing or sealing the system. In some embodiments, the liquid sample absorbed on a sample collector is extracted without a compression mechanism. In some embodiments, the liquid sample absorbed on a sample collector is eluted into a buffer described herein. In some instances, compression of the sample collector in a manner that compresses the sample collector is carried out by the individual from whom the menstrual fluid sample was collected. In some instances, compression of the sample collector is carried out by at a laboratory or other location which processes the sample collector for assaying the collected sample. In some embodiments, the extractor comprises a sample receptacle that receives the sample collector via an opening, and a reservoir that is in fluid communication with the sample receptacle for receiving the biological sample released from the sample collector. In some embodiments, the reservoir and / or receptacle contains a solution comprising one or more reagents for analyzing, preserving, storing, or transporting the collected biological sample. In some instances, placing the sample collector into the first central cavity is carried out by a medical professional, such as an obstetrician or nurse. In some embodiments, the one or more reagents are necessary for hydrolyzing, diffusing, or releasing the biological sample. In some embodiments, the one or more reagents are necessary for analyzing, preserving, or extracting deoxyribonucleic acid, ribonucleic acid, or protein in the biological sample. In some embodiments, the one or more reagents are necessary for reducing analysis background noise. In some embodiments, the one or more reagents are necessary for precipitating or removing a contaminant in the biological sample. In some embodiments, the one or more reagents are necessary for testing the biological sample for a presence or absence of an analyte in the biological sample. In some embodiments, the receptacle contains a reagent that are necessary for dissolving the sample collector upon coming in contact with the sample collector. In some embodiments, accordingly, the sample collector is made of materials that dissolve upon contact with the reagent stored in the receptacle, thereby releasing the biological sample into the reservoir. In some embodiments, the system furthers comprise a cartridge comprising a chamber, wherein the cartridge and / or the chamber is connected to the reservoir via a docking unit, such that upon the cartridge and / or the chamber coming in contact with the reservoir, the released biological sample flows into the cartridge and / or the chamber. In some embodiments, the docking unit comprises a one-way pressure valve. In some embodiments, the docking unit comprises a resealable slit. In some embodiments, the cartridge containing the collected 2020373103   25 Mar 2026 biological sample is covered or sealed. In some embodiments, the cartridge containing the collected biological sample is transported without causing damage or degradation to the collected biological sample. In some embodiments, the system further comprises software or bioinformatics for analyzing the presence of a pathology or disease, and recommending a treatment. In some embodiments, the software is an FDA-approved software. In some embodiments, the system comprises recommending a diet to a subject indicated of nutrition deficiency in the test results, without involving a dietitian or any health care professional. Subjects and prognostic risk factors

[0225] Described herein, in some embodiments, are methods performed to detect a menstrual cycle disorder in a subject. In some embodiments, phenotypic or behavioral characteristics described herein (also referred to as “features ” or “digital biomarkers”) of the subject are utilized to identify a subject at risk for a menstrual cycle disorder. In some embodiments, the features described herein are identified using the survey described herein.

[0226] In some embodiments, a subject is experiencing symptoms of a menstrual cycle disorder such as HMB; AUB; cramping, including cramping which is extreme; fatigue; prolonged menstruation; breast tenderness, which is extreme; migraine; back pain; weight loss; weight gain; one or more irregular cycles; or other symptoms. In some embodiments, the methods described herein are performed on a subject at risk for a menstrual cycle disorder. In some embodiments, an at-risk subject is a subject having HMB, a subject having AUB, a subject previously diagnosed with a menstrual cycle disorder, a subject having a family history of HMB, a subject having a family history of AUB, or a subject having a family history of a menstrual cycle disorder.

[0227] In some embodiments, phenotypic and behavioral characteristics is used to construct a profile of a subject, such as to predict or provide a risk factor of having a menstrual cycle disorder such as HMB or AUB. In some embodiments, inverse probability weighting is applied to adjust for potential confounders or to identify which factors is causative and / or closely associated with having and / or being at risk for a menstrual cycle disorder such as HMB or AUB. In some embodiments, longitudinal data is collected from a subject, such as through one or more surveys to collect phenotypic and behavioral characteristics of the subject. In some embodiments, such surveys are collected a single time, or is collected over days, weeks, months or years. In some embodiments, such surveys are collected from a single subject at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 times over any time period. In some embodiments, such surveys are collected from a single subject not more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 times over any time 2020373103   25 Mar 2026 period. In some embodiments, such surveys are collected from a single subject about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 times over any time period.

[0228] In some embodiments, surveys are collected from a number of different subjects or groups of subjects, and data compared. In some embodiments, survey data from one or more normal subjects (e.g., subjects without a menstrual cycle disorder) is compared with survey data from one or more subjects having HMB and / or AUB, or one or more subjects not having HMB or AUB but having other menstrual cycle disorders. In some embodiments, survey data from one or more subjects is compared with survey data from one or more subjects in the same group (e.g., survey data from a subject having HMB is compared with survey data from other subjects having HMB).

[0229] In some embodiments, survey data is collected and / or compared from at least 1, 5, 10, 50, 100, 500, or 1000 subjects. In some embodiments, survey data is collected and / or compared from not more than 1, 5, 10, 50, 100, 500, or 1000 subjects. In some embodiments, survey data is collected and / or compared from about 1, 5, 10, 50, 100, 500, or 1000 subjects. In some embodiments, survey data is collected and / or compared from between 1 and 1000 subjects, between 1 and 500 subjects, between 1 and 100 subjects, between 1 and 50 subjects, between 1 and 10 subjects, between 1 and 5 subjects, between 5 and 1000 subjects, between 5 and 500 subjects, between 5 and 100 subjects, between 5 and 50 subjects, between 5 and 10 subjects, between 10 and 1000 subjects, between 10 and 500 subjects, between 10 and 100 subjects, between 10 and 50 subjects, between 50 and 1000 subjects, between 50 and 500 subjects, between 50 and 100 subjects, between 100 and 1000 subjects, between 100 and 500 subjects, or between 500 and 1000 subjects. Subjects from whom survey data is collected is in the same group (e.g., normal menstrual bleeding, HMB, AUB, or another group) or in different groups. In some embodiments, a subject is in more than one group at the time of a single survey. In some embodiments, a subject is in one group at the time of a first survey, and in a different group at the time of a second survey. In some embodiments, a subject in a normal menstrual bleeding group is later in an HMB or AUB group, or a subject in an HMB or AUB group is later in a normal menstrual bleeding group.

[0230] In some embodiments, a survey collects data describing one or more phenotypic and / or behavioral characteristics (“digital biomarker”) of subjects. In some embodiments, phenotypic and / or behavioral characteristics of subjects includes characteristics listed in Table 4. In some embodiments, a survey collects data describing a phenotypic or behavioral characteristic 2020373103   25 Mar 2026 (“digital biomarker”) that is not listed in Table 4. In some embodiments, the survey collects the information listed in Figs. 1-23. Table 4: Characteristics of a subject that is utilized to identify a subject at risk for a menstrual cycle disorder CHARACTERISTICS OF SUBJECTS Age location of residence Natural hair color Annual income Level of education Relationship status Frequency of consensual physical touch (outside of sexual contact) Current living situation Religious background Current Religious affiliation Military service Mental health menstruation phenotype pregnancy state number of pregnancies No. and type of tampons or other menstrual products used during period cycle height weight relationship status birth sex race ethnicity sexual orientation gender hormone therapy alcohol consumption cigarette consumption marijuana consumption hemp / CBD consumption exercise diet stress level sleep fertility treatments infertility preeclampsia pre-term labor miscarriage abortion vaginal birth C-section live births mother’s pregnancy history age of first menstruation mother’s age of first menstruation Polycystic Ovarian Syndrome Ovarian cysts Fibroids Breast Cancer Ovarian Cancer Endometrial Cancer Pelvic Inflammatory Disease Chronic or frequent UTIs Ectopic Pregnancy Heart Disease disability Type I Diabetes Type II Diabetes Autoimmune conditions (Lupus, MS, rheumatoid arthritis) Diminished Ovarian Reserve Endometriosis / Adenomyosis family disease history position of uterus Painful periods Heavy bleeding during period Irregular bleeding on or off period Short interval between periods Chronic pelvic pain Lower abdominal or back pain Pain during penetrative sex Pain while defecating, often with cycles of diarrhea and constipation Bloating, nausea, and vomiting Groin pain Pain during exercise vaginal yeast, viral, or bacterial infection vaginal infection treated with antibiotics medication usage freeze preservation of eggs strength of pelvic floor loss of bladder control birth control usage and type duration of hormonal birth control usage sexual history likelihood of orgasm during sex medication or supplement or device usage for sexual enhancement frequency of sex occurrence of sexually transmitted infection abnormal pap smear age of first sexual intercourse number of sexual partners sex during period usage of emergency contraception current menstrual bleeding pattern typical menstrual bleeding pattern cycle length number of days of bleeding 2020373103   25 Mar 2026 consistency of bleeding pattern over menstrual cycles ovulation cycle pain during menstrual window type and level of pain experienced during menstrual window pain outside menstrual window type and level of pain experienced outside menstrual window diet interaction with menstrual-associated pain activity or treatment used for pain relief for menstrual-associated pain spasms in uterus not during menstrual window one-sided, low belly pain during ovulation (mittelschmerz) rectal pain during period or ovulation Number of sanitary products used together for menstrual bleeding passing blood clots during menstruation menstrual bleeding level during sleep menstrual bleeding level during day interference of menstruation with typical daily activities days of heavy flow during menstruation pattern of bleeding during heavy flow days pattern of bleeding during lighter flow days pattern of bleeding throughout menstrual window waist size hip size bust size weight gain potential weight loss potential muscle mass potential to increase or decrease muscle mass areas of fat gain or accumulation body areas having coarse hair Color of menstrual fluid Consistency of menstrual fluid Odor of Menstrual fluid Feel of menstrual fluid Hair loss acne Body hair History of environmental exposure Family history of disease Use of antibiotics Sex education Attitudes toward menstruation Health insurance ER visits Preventative care practices

[0231] In some embodiments, one or more of the characteristics listed in Table 4 is linked to HMB, AUB, or a menstrual cycle disorder. In some embodiments, a characteristic is linked with HMB, AUB, or a menstrual cycle disorder if it correlates with HMB, AUB, or a menstrual cycle disorder (e.g., is present with a disorder more often than a random association), if the characteristic causes HMB, AUB, or a menstrual cycle disorder, or if HMB, AUB, or a menstrual cycle disorder causes the characteristic. In some embodiments, a quantity or level of a characteristic is linked to HMB, AUB, or a menstrual cycle disorder. In some embodiments, a presence or absence of a characteristic is linked to HMB, AUB, or a menstrual cycle disorder. In some embodiments, a link between a characteristic and HMB, AUB, or a menstrual cycle disorder is statistically significant (e.g., p<0.1, p<0.05, or p<0.01).

[0232] In some embodiments, at least 1, at least 2, at least 3, at least 4, at least 5, at least 10, at least 15, at least 20, at least 30, or at least 40 characteristics is linked to HMB, AUB, or a menstrual cycle disorder. In some embodiments, no more than 1, no more than 2, no more than 3, no more than 4, no more than 5, no more than 10, no more than 15, no more than 20, no more than 30, or no more than 40 characteristics is linked to HMB, AUB, or a menstrual cycle disorder. In some embodiments, about 1, about 2, about 3, about 4, about 5, about 10, about 15, about 20, about 30, or about 40 characteristics is linked to HMB, AUB, or a menstrual cycle disorder. In some embodiments, between 1 and 40, between 1 and 30, between 1 and 20, between 1 and 10, between 1 and 5, between 5 and 40, between 5 and 30, between 5 and 20, 80 2020373103   25 Mar 2026 between 5 and 10, between 10 and 40, between 10 and 30, between 10 and 20, between 20 and 40, between 20 and 30, or between 30 and 40 characteristics is linked to HMB, AUB, or a menstrual cycle disorder.

[0233] One or more phenotypic or behavioral characteristics is linked with a specific menstrual cycle disorder, such as endometriosis. In some embodiments, these phenotypic or behavioral characteristics is a subset of the characteristics listed in Table 4. In some embodiments, these phenotypic or behavioral characteristics is an additional characteristic not included in Table 4. In some embodiments, a list of phenotypic and behavioral characteristics that is linked with endometriosis is provided in Table 5. Table 5: Characteristics of subjects which is indicative of Endometriosis CHARACTERISTICS OF SUBJECTS WHICH CAN BE INDICATIVE OF ENDOMETRIOSIS endometriosis or adenomyosis diagnosis surgical diagnosis age of diagnosis years since diagnosis Ovarian endometrioma diagnosis Peritoneal superficial endometriosis diagnosis Deep infiltrating endometriosis diagnosis stage of endometriosis surgical treatment reproductive status removal of endometrial implants retention of ovarian function / loss of reproductive ability removal of uterus and ovaries recurrence Number of surgeries for endometriosis treatment of endometriosis Combination oral contraceptive pills treatment Danazol treatment GnRH agonist treatment Progestins treatment pelvic pain pain killer usage for pelvic pain hormone treatment for pelvic pain pain type pain severity pain frequency activities or treatments that lessen pelvic pain activities or treatments that increase pelvic pain

[0234] In some embodiments, a subject experiencing HMB or AUB experiences HMB or AUB during some or all of their menstrual cycles. In some embodiments, a subject experiencing HMB or AUB experiences HMB or AUB during at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of their menstrual cycles. In some embodiments, a subject experienced HMB or AUB one or more times in the past, is experiencing HMB or AUB for the first time, experience HMB or AUB sporadically, experience HMB or AUB regularly, or experience HMB or AUB during each menstrual cycle.

[0235] In some embodiments, a subject is a human subject. In some embodiments, a subject is a female subject. In some embodiments, the female subject has a chromosomal disorder such as a sex chromosome aberration. In some embodiments, a subject is transgender. In some embodiments, the female subject is experiencing puberty, in her reproductive years, experiencing menopause, or post-menopausal. 2020373103   25 Mar 2026

[0236] In some embodiments, a subject is experienced their first period. In some embodiments, a subject experiences their first period at 9 years of age, 10 years of age, 11 years of age, 12 years of age, 13 years of age, 14 years of age, 15 years of age, 16 years of age, 17 years of age, 18, years of age 19 years of age, or 20 years of age. In some embodiments, some subjects have experienced their first period earlier than 9 years of age or later than 20 years of age. In some embodiments, a subject’s first period has been induced, for example, via a hormone therapy.

[0237] In some embodiments, a risk level is assigned to or calculated for a subject. In some embodiments, a risk level is dependent on the measurement of one or more markers (e.g., expression level of the one or more markers), a phenotypic or behavioral characteristic of the subject, or both. In some embodiments, a risk level of a subject is a quantitative or qualitative assessment of the risk that the subject. In some embodiments, a risk level of a subject indicates a likelihood that the subject has or will develop a condition, such as HMB, AUB, or another menstrual cycle disorder.

[0238] In some embodiments, a subject is stratified into a treatment group based on a risk level assigned to or calculated for them. In some embodiments, stratifying includes assigning a subject to a treatment group or excluding a subject from a treatment group. In some embodiments, stratification is described in further detail below, including in FIGS. 27-29 and in the examples.

[0239] In some embodiments, a reference subject is a subject who is experiencing HMB or AUB. In some embodiments, a reference subject is a subject with a diagnosis of a menstrual cycle disorder or another disorder or disease. In some embodiments, a sample from such a reference subject is a positive control.

[0240] In some embodiments, the subject has a household income of less than $25,000; $25,001 - $50,000; $50,001 - $75,000; $75,001 - $100,000; $100,001 - $150,000; $150,001 - $200,000; or greater than $200,000. In some embodiments, the subject has a highest level of education of no high school, some high school, high school diploma (or equivalent), some college, trade school / technical school certification, associate’s degree, bachelor’s degree, master’s degree, or doctorate or professional degree. In some embodiments, the survey asks the subject to list locations where the subject has lived. Menstrual cycle

[0241] In some embodiments of methods provided herein, the digital biomarker comprises a property of a menstrual cycle. In some embodiments, menstrual fluid or other fluid from the vaginal cavity is collected from a reference subject to provide a reference sample using the 2020373103   25 Mar 2026 methods described herein. In some embodiments, a reference subject is a healthy control subject. In some embodiments, a reference subject experiences normal menstrual bleeding during collection of the menstrual fluid. In some embodiments, a reference subject experiences normal menstrual bleeding during at least 50%, 60%, 70%, 80%, 90%, 95%, 99%, or 100% of their menstrual cycles. In some embodiments, menstrual fluid is collected from more than one reference subject to provide more than one reference sample. In some embodiments, the one or more reference samples serves as control samples. In some embodiments, a reference sample from a reference subject provides a baseline measurement of a property of a fluid sample, such as fluid from the vaginal cavity, such as menstrual fluid, against which a sample from another subject is compared. In some embodiments, the one or more reference samples is used to provide an average or a mean for comparison to one or more other subjects, or to provide a baseline or predetermined threshold for use in comparisons.

[0242] In some embodiments, the survey asks the subject questions about the subject’s menstrual cycle and flow. In some embodiments, the survey asks the subject how long, in days, was the subject’s last cycle (from Day 1 of the subject’s period to the first day of the subject’s next period). In some embodiments, the survey asks a subject that doesn’t use an app or calendar to track the subject’s menstrual cycle to approximate the length of the cycle, including without limitations, noticeably shorter than 28 days, shorter than 28 days but not by much, about 28 days, longer than 28 days but not by much, noticeably longer than 28 days, or don’t know.

[0243] In some embodiments, the survey asks a subject whether during the subject’s cycle, the period start date is completely predictable, somewhat predictable, or not at all predictable. In some embodiments, the survey asks a subject whether during the subject’s cycle, the period end date is completely predictable, somewhat predictable, or not at all predictable. In some embodiments, the survey asks the subject to approximate the frequency of the subject’s period, including without limitations, once a month, more than once a month, every three months, irregular, no identifiable pattern, infrequently, or never. In some embodiments, the survey asks the subject to describe how often the subject changes menstrual products, including without limitations, every hour, every 2 hours, every 4 hours, every 6 hours, every 8 hours, or every 10 hours. In some embodiments, the subject had a menstrual cycle during the past month. In some embodiments, the subject had most recent menstrual bleeding that was typical of what is normal for the subject. In some embodiments, the subject did not have a menstrual cycle due to birth control, pregnancy, or being perimenopausal. In some embodiments, the subject had bleeding when not on the subject’s period. 2020373103   25 Mar 2026

[0244] In some embodiments, information on a typical menstrual cycle comprises the length of a menstrual cycle, the number of days of bleeding of a menstrual cycle, the frequency of a menstrual cycle. In some embodiments, the color of the menstrual fluid is black, brown, dark red, purple maroon, bright red, pink, orange, yellow, or gray, as depicted in FIG. 2. In some embodiments, the texture of the menstrual fluid is classified as runny liquid, thick consistency, clots / thick chunks, slippery with mucus, stringy, or a combination thereof. In some embodiments, the odor of the menstrual fluid is metallic / coppery, rotten, fishy, like onions, gym sweat, tangy / fermented, or sweet. In some embodiments, the feel of the menstrual flow is a drip, a trickle, gushing, a flood, or nothing. In some embodiments, the survey asks the subject to describe their period as spotting, very light, light, moderate, or heavy. An example of this is seen in FIG. 1. In some embodiments, the survey asks the subject how many days (including spotting) did the subject bleed for during the subject’s period. In some embodiments, the survey asks the subject to chronologically list the sequence of spotting, light flow days, medium flow days and heavy flow days the subject experiences during menstruation. In some embodiments, the survey asks the subject if the subject’s menstrual flow (heaviness) is nearly identical, somewhat similar, or completely different month to month.

[0245] In some embodiments, the survey asks a subject how many times the subject bleeds through an outfit (not underwear) during a heavy flow, a medium flow, or a light flow, even when using typical menstrual products. In some embodiments, the survey asks a subject how many times the subject gets up, after the subject has gone to bed, to change menstrual products during a heavy flow, a medium flow, or a light flow. In some embodiments, the survey asks the subject how many light, regular, super, super plus, and ultra tampons the subject fully soaks on a given day (24 hours). In some embodiments, the survey asks the subject how many light, regular, super, super plus, and ultra tampons the subject fully soaks on a heavy flow, a medium flow, or a light flow day during the daytime and nighttime. An example of tampons sizes is seen in FIG. 3. In some embodiments, the survey asks the subject how many pantyliner, ultra-thin, regular, maxi / super, overnight, and post-partum pads the subject fully soaks on a heavy flow, a medium flow, or a light flow day (24 hours). In some embodiments, the survey asks the subject how many pantyliner, ultra-thin, regular, maxi / super, overnight, and post-partum pads the subject fully soaks on a heavy flow, a medium flow, or a light flow day during the daytime and nighttime. In some embodiments, an example of pad sizes is seen in FIG. 4. In some embodiments, the survey asks the subject to estimate how many times the subject empties or changes a menstrual cup, menstrual disk, period panties, or adult diaper per heavy flow, medium 2020373103   25 Mar 2026 flow, or light flow day (24 hours). In some embodiments, the survey asks the subject to estimate how full the subject’s menstrual cup, menstrual disk, period panties, or adult diaper was when changed. In some embodiments, these products are shown in FIG. 5.

[0246] In some embodiments, the survey collects information on how many menstrual products are fully used. In some embodiments, the survey collects information on how many menstrual products are used on a heavy flow day, a medium flow day, or a light flow day. In some embodiments, a survey collects information on how many products are used during the daytime or during the nighttime. In some embodiments, the menstrual products is a tampon, a pad, a menstrual cup, a menstrual disc, period panties, or an adult diaper. In some embodiments, the tampons is light, regular, super, super plus, or ultra tampons. In some embodiments, the pad is a panty-liner, an ultra-thing pad, a regular pad, a maxi / super pad, an overnight pad, or a postpartum pad.

[0247] In some embodiments, the survey collects information about menstrual product use. In some embodiments, the survey collects information about current use or past use of menstrual products. In some embodiments, the menstrual product is organic tampons, nonorganic tampons, organic pads, nonorganic pads, menstrual cup, menstrual disc, period panties, adult diaper, or bladder control pads. In some embodiments, the survey asks the subject which of the following menstrual products the subject currently uses, including without limitations, organic tampons, nonorganic tampons, organic pads, nonorganic pads, menstrual cup, menstrual disc, period panties, adult diaper, or bladder control pads. In some embodiments, the survey asks the subject which of the following menstrual products the subject has ever used, including without limitations, organic tampons, nonorganic tampons, organic pads, nonorganic pads, menstrual cup, menstrual disc, period panties, adult diaper, or bladder control pads.

[0248] In some embodiments, the survey asks the subject how many years the subject has used organic tampons. In some embodiments, the survey asks the subject how many years the subject has used nonorganic tampons. In some embodiments, the survey asks the subject how many years the subject has used organic pads. In some embodiments, the survey asks the subject how many years the subject has used nonorganic pads. In some embodiments, the survey asks the subject how many years the subject has used a menstrual cup. In some embodiments, the survey asks the subject how many years the subject has used a menstrual disk. In some embodiments, the survey asks the subject how many years the subject has used period panties. In some embodiments, the survey asks the subject how many years the subject has used adult diapers. In 2020373103   25 Mar 2026 some embodiments, the survey asks the subject how many years the subject has used bladder control pads.

[0249] In some embodiments, the subject has a pre-existing condition / disability or physical limitation that influences the products the subject uses during the subject’s period. In some embodiments, the survey asks the subject to explain why and how the subject manages the subject’s menses. In some embodiments, the survey asks if religious or cultural norms have influenced the subject’s menstrual management routine and to explain how.

[0250] In some embodiments, the survey collects information on cycle tracking. In some embodiments, the survey collects information on ovulation. In some embodiments, the survey collects information on methods of tracking ovulation. In some embodiments, the methods are spotting, mittelschmerz (lower belly pain, usually one-sided), LH urine test, PdG urine test, testing consistency of cervico-vaginal fluid, or, basal body temperature. In some embodiments, the survey collects information about methods of tracking a period. In some embodiments, the subject tracks aspects of the subject’s menstrual cycle and / or ovulation using a paper calendar, a mobile app, or another method. If the subject tracks aspects of the subject’s menstrual cycle and / or ovulation using a mobile app, the survey asks the subject which mobile app(s) the subject uses and why the subject likes one app more than another. If the subject tracks aspects of the subject’s menstrual cycle and / or ovulation using a mobile app, the survey asks how accurate the mobile app is in predicting the start date of the subject’s period, with the options being within 1, 2, 3, 4, 5, 6, ,7 ,8 ,9, or 10 days of the start date of the subject’s period.

[0251] In some embodiments, the survey collects information on missed days of work or school, reduced work capacity, impact on life, or impact on career due to menstruation. In some embodiments, the survey asks a subject how many days of work or school the subject misses due to a typical period. In some embodiments, the survey asks a subject how many days the subject shows up to work or school but is unable to be fully present or perform to full capacity due to a typical period. In some embodiments, the survey asks a subject to rate how much the subject’s period has affected the subject’s life or career on a scale of 1 to 10 (1 = no impact, 10 = failed classes or lost promotions due to period). In some embodiments, a subject experiences pain during menstruation. In some embodiments, menstrual pain is rated between 0 and 10 on the Mankoski pain scale.

[0252] In some embodiments, the survey collects information on menstrual clots. In some embodiments, the survey asks, during a typical period, how many times does the subject notice menstrual clots (blobs of blood, tissue, and mucus which can look gel-like or clumpy) whether 2020373103   25 Mar 2026 it’s seeing it or feeling it. If the subject experiences menstrual clots, the survey asks the subject to describe the size of the clots, including without limitations, smaller than a dime, size of a dime, size of a nickel, size of a quarter, or size of a ping-pong ball.

[0253] In some embodiments, the survey collects information on habits during menstruation. In some embodiments, the survey asks if finances affect how a subject manages the subject’s period. In some embodiments, if finances do affect how a subject manages a period, the survey asks the subject to rate the following statements on a scale of 1 to 10 (1 being “I can’t relate to this statement” and 10 being “this statement strongly resonates with me”): The survey asks if during a period, the subject wears specific clothing to hide or minimize leaks or packs a change of clothes in case of leaks. In some embodiments, the survey asks if as the start date of the subject’s period approaches, the subject starts carrying spare menstrual products; pre-emptively changes their diet or exercise routine; pre-emptively takes pain medication in anticipation of pain; starts avoiding essential activities like grocery shopping; starts avoiding leisure activities like hanging out with friends; or reschedules appointments. In some embodiments, the survey asks a subject how often the subject checks the back of a seat or clothing for leaks during a typical period.

[0254] In some embodiments, a subject is experience symptoms either chronically, consistently, or occasionally. In some embodiments, symptoms include painful period, heavy bleeding during a period, irregular bleeding on or off a period, short interval between periods, chronic pelvic pain, lower abdominal or back pain, pain during penetrative sexual intercourse, pain during defecation (including normal defecation, diarrhea, and constipation), bloating, nausea, vomiting, groin pain, pain during exercise, migraine, fatigue, throbbing veins in legs, shooting rectal pain, acne, pale skin, increased body odor, or greasy hair. In some embodiments, a subject experiences spasms in their uterus when menstruating and / or when not menstruating. In some embodiments, a subject experiences one-sided, low belly pain during ovulation, i.e., mittelschmerz. In some embodiments, a patient experiences shooting rectal pain during a menstrual cycle or during ovulation. In some embodiments, the survey asks the subject to describe the role pain plays in the subject’s life.

[0255] The survey asks which of the following helps relieve subject’s menstrual pain: pain medication (over the counter), pain medication (prescription), cannabis, ice, kava, hot bath, black cohosh, emptying bladder, relaxation, music, heating pad, meditation, sex / orgasm, acupuncture, lying down, massage, bowel movement, laxatives / enema, TENS unit (electrical stimulus), exercise, or other. In some embodiments, the survey asks the subject if eating sugar, 2020373103   25 Mar 2026 animal products, alcohol, junk food, spicy food, salty food, or any other foods increases menstrual pain. In some embodiments, the survey asks the subject if eating salmon, banana, dark chocolate, citrus, spicy foods, dark leafy vegetables, watermelon, chamomile, herbal teas, or other foods helps relieve menstrual pain.

[0256] In some embodiments, the survey collects information on attitudes towards menstruation. In some embodiments, the survey collects information on the age in which a subject first learned about menstruation, how a subject first learned about menstruation, childhood communication on menstruation, shame around menstruation, influences on thoughts on menstruation, or availability of menstrual products. In some embodiments, the survey asks how old a subject was when the subject first learned about menstruation. In some embodiments, the subject is first learned about menstruation from, including without limitations, parents / family, friends / peers, puberty & self-help books, online resources, or other resources. In some embodiments, the survey asks a subject to rate how openly the subject’s family communicated about periods when growing up on a scale of 1-10 (1 = no discussions of menstruation, hiding menstrual products and waste; 10 = encouraging and celebrating menstruation, menstrual products stored openly). In some embodiments, the survey asks what kinds of messages the subject received around menstruation while growing up. In some embodiments, the subject regularly hid period products from view when going to the bathroom in public in the past. In some embodiments, the subject currently hides period products from view when going to the bathroom in public. In some embodiments, the survey asks if a subject has ever felt shame around the subject’s period. If the subject has felt shame in the past, the survey asks the subject to rate the shame on a scale of1 -10. In some embodiments, the survey may also ask the subject to rate the shame the subject currently feels on a scale of 1 - 10. In some embodiments, the survey asks the substitute to identify the major influences that shaped the subject’s current views on menstruation, including without limitations, family, friends / peers, religious beliefs, cultural norms, menstruation advocacy groups or nonprofits, entertainment media (e.g. books, movies, TV shows), social media (e.g. influencers, threads on social networks), or other.

[0257] In some embodiments, the survey asks a subject to describe the availability of menstrual products in bathrooms outside the subject’s home on a scale of 1-10. In some embodiments, the survey asks if a subject’s school or workplace provides any of the following period-related accommodations: paid menstrual leave, ability to work from home during the subject’s period, menstrual products available in bathrooms, freely available pain management products, such as pain medication or heating pads, or any other accommodations. 2020373103   25 Mar 2026

[0258] In some embodiments, a subject has a uterus which is anteverted (i.e., tipped forward or aimed toward the belly), retroverted (i.e., tipped backwards, or aimed toward the rectum), anteflexed (i.e., top of uterus points forward relative to cervix while front of uterus is concave), or retroflexed (i.e., top of uterus points backward relative to cervix while front of uterus is convex). In some embodiments, the survey asks the subject to indicate the position of the subject’s uterus using FIG. 19 as a reference. Menstrual Cycle Disorders and autoimmune disorders

[0259] In some embodiments of methods herein, the digital biomarker comprises a property related to a menstrual cycle disorder or other autoimmune disorder. In some embodiments, a subject has a menstrual cycle disease or disorder, or other disorder which presents with, or cause, HMB or AUB. In some embodiments, diseases and disorders that a subject has include eating disorders; extreme weight loss; excessive exercise; polycystic ovary syndrome (PCOS); ovarian cysts; premature ovarian failure; breast cancer; ovarian cancer; infertility; diminished ovarian reserve; chronic or frequent urinary tract infections; ectopic pregnancy; heart disease; type 1 diabetes; type 2 diabetes; an autoimmune condition such as lupus, multiple sclerosis, or rheumatoid arthritis; pelvic inflammatory disease (PID); endometriosis; fibroids (e.g., uterine fibroids); adenomyosis; cervical cancer; endometrial cancer; uterine cancer; or infection of the cervix or endometrium. In some embodiments, HMB or AUB accompanies a disease affecting the kidney, liver, thyroid, or adrenal glands.

[0260] In some embodiments, the survey collects information on diagnosis with endometriosis or adenomyosis, age of diagnosis, tests involved in diagnosis, subtypes of endometriosis, stage of endometriosis, medication to treat endometriosis, surgical treatment to endometriosis, reoccurrence of endometriosis, pain associated with endometriosis, pain relief strategy, or pelvic pain. In some embodiments, a subject has endometriosis. In some embodiments, the subject has been surgically diagnosed with endometriosis or adenomyosis. In some embodiments, the subject, has been diagnosed with adenomyosis by either ultrasound or MRI. In some embodiments, the subject is suspected of having endometriosis or adenomyosis. In some embodiments, the survey asks subjects that have been surgically diagnosed with either endometriosis or adenomyosis by a doctor how old the subjects were when diagnosed and how many years has the subject lived with endometriosis. In some embodiments, the survey may also ask what tests the doctor used to diagnosis the subject, which could include ultrasound, pelvic exam, surgery, or other tests. 2020373103   25 Mar 2026

[0261] Endometriosis is a condition when endometrium grows on other pelvic organs. In some cases, types of endometriosis include ovarian endometrioma, peritoneal superficial endometriosis, deep infiltrating endometriosis, or adenomyosis. In some embodiments, endometriosis is stage 1 (minimal), stage 2 (mild), stage 3 (moderate), or stage 4 (severe). In some embodiments, a subject had endometriosis for at least 1 month, 6 months, 1 year, 2 years, 3 years, 4 years, 5 years, or more. In some embodiments, a subject has had surgery for endometriosis. In some embodiments, the surgery is conservative, semiconservative, or radical. In some embodiments, the medication is combination oral contraceptive pills (COCs), Danazol, GnRH agonist, or progestins.

[0262] In some embodiments, a subject has adenomyosis. Adenomyosis is a condition where the endometrium breaks through or invades the muscle wall of the uterus. In some cases, symptoms of adenomyosis include menstrual cramps, lower abdominal pressure, bloating before periods, or HMB. In some embodiments, a subject has had adenomyosis for at least 1 month, 6 months, 1 year, 2 years, 3 years, 4 years, 5 years, or more.

[0263] In some embodiments, the survey collects information on fibroids. In some embodiments, the information is age at diagnosis, years since diagnosis, method of diagnosis, type of fibroids, surgical treatment, age at treatment, reoccurrence at treatment, medication to treat fibroids, pelvic pain, pain relief, and frequency of pelvic pain. in some embodiments, the type of fibroid is intramural (in the uterine wall), subserosal (on the outside of the uterine wall), submucosal (under the lining of the uterine cavity), or pedunculated (a stalk like growth that can attach to many places). In some embodiments, the surgical treatment is myomectomy (laparoscopic / open / hysteroscopic), laparoscopic myomectomy, open myomectomy, hysteroscopic myomectomy, hysterectomy, uterine fibroid embolization (ufe), endometrial ablation, myolysis, partial hysterectomy, MRI-guided focused ultrasound surgery, watching / wait and see / waiting for menopause, or complementary and / or alternative medicine (CAM). In some embodiments, the medication is combination oral contraceptive pills (COCs), GnRH agonist, Progestins, Tranexamic acid (Lysteda, Cyklokapron), or Ulipristal acetate.

[0264] In some embodiments, the survey asks how much pelvic pain the subject had during the subject’s last period, including without limitations, no pain, mild cramps (medication never or rarely needed to continue daily activities), moderate cramps (medication usually needed to continue daily activities), or severe cramps (medication and bed rest needed to continue daily activities). In some embodiments, the survey asks how often the subject had pelvic pain during the subject’s period, including without limitations, seldom (less than a quarter of my periods), 2020373103   25 Mar 2026 often (a quarter to half of my periods), usually (more than half of my periods), or always (every period).In some embodiments, the subject is taken pain-killers or hormones for pelvic pain during the subject’s last period. In some embodiments, the painkillers may be prescribed by a doctor, over-the-counter (e.g. aspirin, ibuprofen, paracetamol / acetaminophen, naproxen), hormones, or a cannabinoid based product.

[0265] The survey asks if during the subject’s last period, the pelvic pain prevented the subject from going to work or school or carrying out daily activities (even if taking pain-killers). In some embodiments, the survey asks the subject to rate how severe the subject’s pelvic pain was at its worst during the subject’s last period from 0-10 on the Mankoski Pain Scale.

[0266] Th subject may have had pelvic pain within the last 3 months that felt, including without limitations, throbbing, shooting, stabbing, sharp, cramping, gnawing, hot-burning, aching, heavy, tender, splitting, tiring-exhausting, sickening, fearful, punishing-cruel, or other. In some embodiments, the survey asks the subject to identify factors that makes the pelvic pain worse such as sitting, full bladder or urinating, time of day, stress, bowel movement, constipation, full meal, intercourse or orgasm, standing or walking, exercise, weather, contact with clothing, or coughing / sneezing. In some embodiments, the survey asks what helps the subject’s pelvic pain, including without limitations, prescription pain medication, over the counter pain medication, relaxation, lying down, music, massage, ice, heating pad, bowel movement, hot bath, meditation, laxatives / enema, emptying bladder, cannabinoid based products, herbal remedies, black cohosh, kava, or other.

[0267] In some embodiments, the survey collects information on an autoimmune disease. In some embodiments, the autoimmune disease is Alopecia areata, Antiphospholipid antibody syndrome (APL), Autoimmune hepatitis, Celiac disease, Crohn’s disease, Diabetes type 1, Graves’ disease, Guillain-Barre syndrome, Hashimoto’s disease, Hemolytic anemia, Idiopathic thrombocytopenic purpura (ITP), Inflammatory bowel disease (IBD), Inflammatory myopathies, Multiple sclerosis (MS), Myasthenia gravis (MG), Primary biliary cirrhosis, Psoriasis, Rheumatoid arthritis, Scleroderma, Sjogren’s syndrome, Systemic lupus erythematosus, Vitiligo, or a combination thereof. In some embodiments, the survey collects information on age of first diagnosis, years since diagnosis, or time to diagnosis. In some embodiments, the survey collects information on medications or complementary or alternative medicine used to treat an autoimmune disorder. In some embodiments, the medication is Corticosteroids (e.g. Prednisone); NSAIDS (e.g. Advil); DMARDs (e.g. Methotrexate, Plaquenil); Janus kinase inhibitor (e.g. Xeljanz); Calcineurin inhibitor (e.g. Neoral, Astagraf XL); mTOR inhibitor (e.g. 2020373103   25 Mar 2026 Rapamune, Afinitor); IMDH inhibitor (e.g. Imuran, CellCept); DMTs (e.g. Lemtrada, Aubagio); Hormones (e.g. Levothyroxine); Biologic (e.g. Humira, Rituxan, Stelara); Monoclonal antibodies (e.g. Simulect, Zinbryta); IVIG (e.g. Immunoglobulin therapy); Plasmapheresis; surgery; or other treatments. In some embodiments, the complementary or alternative medicine is vitamins and supplements (i.e. Vitamin D, probiotics), yoga / exercise, acupuncture, diet, elimination of toxins, functional medicine, nutritionist, physical therapist, or a combination thereof. In some embodiments, the survey collects information on reducing flareups, managing symptoms, a relationship between menstruation and an autoimmune disorder, or impact on quality of life.

[0268] In some embodiments, the survey asks what symptoms a subject experiences during a flare-up, including without limitations, fatigue, joint pain, digestive issues, dermatological issues (i.e. rash, irritation), pain, hair loss, weight loss, weight gain, muscle weakness, brain fog, anemia, numbness and tingling, swollen glands, or other symptoms. In some embodiments, the survey asks how a subject knows a flare up is imminent, including without limitations, unknown, stress, eating a “trigger” food, a major life change, overdoing it, not taking prescribed medicine, change in weather, change in seasons, bacterial or viral infection, or lack of sleep.

[0269] In some embodiments, the subject believes there is a correlation between the subject’s period and symptoms / flares of the autoimmune disease(s). In some embodiments, the subject’s period is impacted during an autoimmune disease(s) flare up, including without limitations, heavier flow than normal, lighter flow than normal, irregular or increased frequency of cramping, increased intensity of cramping, or increase in clotting. In some embodiments, the survey asks how a subject’s autoimmune disease(s) is impacted when the subject has their period, including without limitations, more likely to experience a flare, increased fatigue, increased pain, autoimmune symptoms worsen, or autoimmune symptoms improve. In some embodiments, the survey asks if during the subject’s last period, the symptoms (either period related or autoimmune related) prevented the subject from going to work or school or carrying out daily activities. In some embodiments, the subject’s ability to work full-time, ability to work part-time, ability to attend school, social life, romantic life, recreational travel, parenting, hobbies, or exercise is impacted by the subject’s autoimmune disease greatly, somewhat, or not at all.

[0270] In some embodiments, the subject has been or is currently pregnant. In some embodiments, if the subject has been or is pregnant, the subject’s autoimmune disease(s) was impacted during pregnancy, including without limitations, autoimmune disease(s) went into a 2020373103   25 Mar 2026 non-drug-induced remission, experienced a flare of autoimmune disease(s), autoimmune symptoms improved, autoimmune symptoms worsened, or other. In some embodiments, the subject has been on treatment prior to getting pregnant, and the survey asks if the subject stayed on this treatment regimen through the pregnancy. In some embodiments, the subject’s autoimmune diseases have been impacted post-partum, including without limitations, autoimmune disease(s) went into a non-drug-induced remission, experienced a flare of autoimmune disease(s), autoimmune symptoms improved, autoimmune symptoms worsened, or other. Sexual Behaviors and Education

[0271] In some embodiments of methods provided herein, the digital biomarker comprises a property related to a sexual behavior or education of the subject as described herein. In some embodiments, the survey collects information on sexual education. In some embodiments, the sexual education is abstinence, consent, how to say “no” to sex, different birth control methods, STIs, how to use a condom, how to prevent HIV, HPV vaccine, or a combination thereof. In some embodiments, the sexual education is formal or informal. In some embodiments, the survey collects information about the age of sexual education or the source of sexual education. In some embodiments, the survey collects information about communication about sex.

[0272] In some embodiments, the survey asks the subject to identify the sex-ed topics for which the subject received “formal” or informal instruction before the age of 18, including without limitations, abstinence, consent, how to say “no” to sex, different birth control methods, STIs, how to use a condom, how to prevent HIV, or the HPV vaccine. In some embodiments, if the subject received formal sex-ed instruction, the subject received it prior to the subject’s first experience of intercourse.

[0273] In some embodiments, the survey asks the subject to identify the sex-ed topics for which the subject received “informal” instruction (from parents or guardians) before the age of 18, including without limitations, abstinence, consent, how to say “no” to sex, different birth control methods, STIs, how to use a condom, how to prevent HIV, or the HPV vaccine. If the subject received informal sex-ed instruction. In some embodiments, the subject is received it prior to the subject’s first experience of intercourse.

[0274] In some embodiments, the survey asks which sources of sex education the subject received outside of school, including without limitations, puberty & self-help books, bird & the bees conversation with my parents, friends and siblings, pornographic videos & photos, romance novels, or other. 2020373103   25 Mar 2026

[0275] In some embodiments, the subject speak with the subject’s partner about sex, sexual needs and desires often, when necessary, or never. In some embodiments, the subject speaks with friends about the subject’s sex life often, when necessary, or never.

[0276] In some embodiments, the survey collects information on being sexually active, frequency of sex, STIs, age of first vaginal intercourse, sex during a period, emergency contraception, sexual pleasure, masturbation, orgasm, orgasm supplements, sex toys, erotic literature, or other materials to enhance sexual arousal. In some embodiments, the subject has been diagnosed with dyspareunia (painful sexual intercourse) or experienced genital pain just before, during or after sexual intercourse. In some embodiments, the subject has been treated for a sexually transmitted infection. In some embodiments, the survey asks how old the subject was during the subject’s first sexual encounter. In some embodiments, the survey asks how old the subject was when the subject first had vaginal intercourse.

[0277] In some embodiments, a subject is sexually active. In some embodiments, a subject has sexual intercourse less than 10 times per year, about 10 times per year, about 1-3 times per month, about 1-3 times per week, or more than 3 times per week. In some embodiments, a subject had their first sexual encounter (e.g., sexual intercourse) at age 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, older than age 25, or younger than age 13. In some embodiments, a subject has sexual intercourse during their period or not have sexual intercourse during their period.

[0278] In some embodiments, the survey asks the subject what frequency best describes how often the subject receives welcome and consensual physical touch (outside of sexual contact), including without limitations, rarely, occasionally, regularly, or frequently.

[0279] In some embodiments, a subject had 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more sexual partners during their lifetime. In some embodiments, a subject had 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more sexual partners at the time a sample is collected.

[0280] In some embodiments, the subject has sex during menstruation. If the subject does not have sex during menstruation, the survey asks if this is due to the subject’s preference or the preference of the subject’s partner. In some embodiments, the survey asks how many times the subject has used emergency contraception (Plan B, Ella, etc). In some embodiments, the subject knows that Plan B is ordered through Amazon.

[0281] In some embodiments, the survey asks about a subject’s views on masturbation. In some embodiments, the survey asks if the subject typically reaches orgasm while masturbating; if the subject has ever had an orgasm with a sexual partner; or if the subject has ever taken over-the- 2020373103   25 Mar 2026 counter medications or supplements to enhance orgasm and which medications or supplements were taken; if the subject uses “toys” (such as vibrators) to achieve sexual pleasure; if the subject uses visual material (pictures / videos) to enhance sexual arousal; if the subject uses aural material to enhance sexual arousal (music, erotic podcasts); if the subject uses erotic literature to enhance sexual arousal; if the subject uses any apps to improve the subject’s sex life, and if so, the survey asks what apps the subject uses.

[0282] In some embodiments, a subject has a sexually transmitted infection or has had a sexually transmitted infection in the past. In some embodiments, a sexually transmitted infection is bacterial vaginosis, chlamydia, gonorrhea, genital herpes, hepatitis, human immunodeficiency virus, acquired immunodeficiency syndrome, human papillomavirus, pelvic inflammatory disease, syphilis, or trichomoniasis. Pregnancy and Childbirth

[0283] In some embodiments of methods provided herein, the digital biomarker is a property related to pregnancy, fertility, and childbirth of the subject. In some embodiments, the survey collets information about fertility. In some embodiments, the survey collects information on attempts to get pregnant, time to get pregnant, fertility treatment, infertility, egg freezing, egg donor, gestational carrier, embryo freezing, pregnancy, pre-term labor, pre-eclampsia, miscarriage, abortion, or frequency of medical care during pregnancy. In some embodiments, the survey collects information on giving birth, number of times giving birth, C-sections, vaginal delivery, breastfeeding, formula feeling, mothers age at birth of first child, mother’s age at birth of last child, or a combination thereof. In some embodiments, the survey collects information about medical treatment after birth, time off work after birth before returning to work, childcare after birth, stiches after birth, postpartum hemorrhaging, postpartum bleeding, passing small clots, passing clots bigger than the size of a quarter, headaches, high blood pressure, swelling in hands and feet, blurred vision, sudden weight gain, leg pain, pain that felt like a pulled muscle, swelling in legs, painful urination, difficulty urinating, constipation, chills or fever, vaginal discharge with no odor, vaginal discharge with odor, heart palpitations, chest pain, difficulty breathing, afterpains (abdominal pains which can be especially feel while breastfeeding), increasingly worsening lower belly pain, back pain, incontinence, soreness in the perineal and / or vaginal area, anxiety, stress, depression, obsessive-compulsive behavior, emotional instability, sadness, diastasis recti (a gap between the right and left abdominal wall muscles which can give the appearance of a pregnant belly months after birth), postpartum preeclampsia, stroke, deep vein thrombosis, c-section infection, episiotomy infection, hemorrhoids, baby blues 2020373103   25 Mar 2026 (overwhelmed, fragile, sad for a few weeks after birth), postpartum depression (mood swings, sadness, insomnia, irritability, suicidal thoughts, inability to care for new born that lasts longer than a few weeks), lactation consultant, experiences breastfeeding, pelvic therapy, sex drive, weight gain, changes in shoe size, hair loss, stretch marks, return of period after pregnancy, or a combination thereof. In some embodiments, the subject is experienced postpartum hemorrhaging to an extent that the subject filled more than one pad every hour.

[0284] In some embodiments, a subject is pregnant. In some embodiments, the pregnancy is a normal pregnancy or a complicated pregnancy. In some embodiments, complications comprise pre-eclampsia, eclampsia, stillbirth, a history of stillbirth, miscarriage, a history of miscarriage, high blood pressure, gestational diabetes, preterm labor, nausea and vomiting, hyperemesis gravidarum, or iron deficiency anemia. In some embodiments, a pregnant subject is at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, or more than 40 weeks pregnant.

[0285] In some embodiments, a subject is a post-partum subject. In some embodiments, a postpartum subject is at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or more weeks post-partum.

[0286] In some embodiments, the survey asks if a subject has ever given birth. If the subject has given birth, the survey asks how many times the subject has given birth and the age at which the subject first gave birth. In some embodiments, the survey asks if a subject has ever had a C-section or vaginal delivery. In some embodiments, a subject has a baby. In some embodiments, a baby is 0 months, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 12 months, 18 months, or 24 months of age.

[0287] In some embodiments, a subject has been pregnant before. In some embodiments, a subject has had 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more pregnancies. In some embodiments, a subject has had their first pregnancy at age 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, or 40. In some embodiments, a subject has had their first pregnancy earlier than age 13 or later than age 40. In some embodiments, a subject has had 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more live births. In some embodiments, a subject has had 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more stillbirths. In some embodiments, a subject has had 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more miscarriages. In some embodiments, a birth (e.g., a live birth or a still birth) is via a Cesarean section or a vaginal delivery.

[0288] In some embodiments, the survey asks how often the subject saw a doctor during pregnancy, including without limitations, once a month until week 28, then once a month until week 36, then every week in the last month of pregnancy; once a trimester; once before birth; at 2020373103   25 Mar 2026 birth; or other. In some embodiments, the survey asks how long after giving birth was the first time the subject saw a doctor again, including without limitations, within 3 weeks of birth for a normal checkup, for a 6 week normal checkup, or before a normal scheduled checkup for an emergency. In some embodiments, the subject has taken leave for 1 week, 2 weeks, 1 month, 2 months, 3 months, 4 to 6 months, 6 to 12 months, or longer than 12 months before returning to work. In some embodiments, the subject does not return to work.

[0289] In some embodiments, a subject is having or has a history of miscarriage. In some embodiments, a subject has had a miscarriage in the past 1, 2, 3, 4, 5, 6, 7, 8, 9 10, 11, or 12 months. In some embodiments, a subject has had a miscarriage in the past 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 years. In some embodiments, a subject has had multiple miscarriages. Some subjects has a family history of miscarriage. If the subject has had one or more miscarriages, the survey asks how many weeks pregnant the subject was during the last miscarriage and how long it took for the subject’s period to return after the last miscarriage.

[0290] In some embodiments, a subject has had an abortion in the past. In some embodiments, a subject has had more than one abortion. In some embodiments, a subject has had 0, 1, 2, 3, 4, or more abortions. If the subject has had one or more abortions, the survey asks how many weeks pregnant the subject was during the last abortion and how long it took for the subject’s period to return after the last abortion.

[0291] In some embodiments, a subject has frozen their eggs. In some embodiments, a subject has gone through 1, 2, 3, 4, or more cycles of egg freezing. In some embodiments, a subject is frozen their embryos. In some embodiments, a subject has gone through 1, 2, 3, 4, or more cycles of egg freezing. In some embodiments, the subject has been an egg donor or gestational carrier for another individual or couple. In some embodiments, the subject is frozen the subject’s embryos. If so, the survey asks how many cycles the subject has gone through.

[0292] In some embodiments, a subject is exclusively breastfeeding their baby, mostly breastfeed their baby, breastfeed their baby about half the time, breastfeed their baby occasionally, or never breastfeed their baby. In some embodiments, the survey asks how many months the subject pursed this newborn feeding strategy. In some embodiments, the subject has had a partner to help with nighttime feedings or wakings. In some embodiments, the survey asks if the subject had extended family, friends or nanny care to help with baby care and household care after birth. In some embodiments, the subject had help for at least about 2 weeks, 1 month, 2 months, 3 months, 4 to 6 months, 6 to 12 months, or longer than 12 months. 2020373103   25 Mar 2026

[0293] In some embodiments, the subject used a lactation consultant. In some embodiments, the survey asks the subject to rate how stressful the subject found the experience of getting the newborn to latch on a scale of 1-10. In some embodiments, the survey asks the subject to rate how frustrating the subject found the experience of nursing to be the first time. In some embodiments, the subject is experienced sore nipples and breasts after birth, and if so, the survey asks how many weeks the soreness lasted after the subject first started breast feeding. In some embodiments, the survey asks if the soreness resolved after the subject’s baby latched correctly. In some embodiments, the subject is developed painful cracks on the nipples, mastitis (inflammation of the breasts), or breast abscesses (painful build-up of pus in breasts). In some embodiments, the subject has seen a pelvic therapist during pregnancy to prepare for giving birth or after birth for pelvic massage, back pain, incontinence or other pregnancy / postpartum issues,

[0294] In some embodiments, the survey asks which of the following items the subject used in postpartum care: ibuprofen / acetaminophen, sitz bath, perineal spray, healing foam, maxi pads, upside down squirt bottle to wash perineal area, postpartum underwear, ice packs for vaginal / perineal area, kegels, nipple balm, or other. In some embodiments, the subject is found it easy to learn about the over the counter products (such as postpartum underwear, “MomWasher” peri bottles) which were most useful in the subject’s postpartum care. In some embodiments, the survey asks from whom did the subject find out about the most useful products to use during postpartum care, including without limitations, doctor, nurse, lactation consultant, pregnancy preparation classes, family and friends, books, pregnancy / childbirth / parent Facebook groups, Instagram accounts, Googling, or other. In some embodiments, the subject is bought most products for postpartum care at big box retailers (Target, Walmart), pharmacies (CVS, Walgreens etc.), Amazon, independent vendors specializing in baby care with store fronts, or independent vendors specializing in baby care on-line.

[0295] In some embodiments, the survey asks how long it took after childbirth for the subject’s sex drive to return, including without limitations, within a month, 1 to 3 months, 3 to 6 months, 6 to 12 months, or longer than 12 months. In some embodiments, the survey asks how much weight the subject gained during pregnancy and if the subject returned to a pre-pregnancy weight after giving birth. If the subject did return to the subject’s pre-pregnancy weight, the survey asks the subject to rate on a scale of 1 to 10, 1 being easy and 10 being very difficult, how hard it was to return to a pre-pregnancy weight. In some embodiments, the survey asks how long it took for the subject to return to a pre-pregnancy weight. In some embodiments, the subject experienced an increase in shoe size during pregnancy, and if so, the survey asks how 2020373103   25 Mar 2026 long it took the subject to return to the subject’s pre-pregnancy shoe size, including without limitations, within a month, 1 to 3 months, 3 to 6 months, 6 to 12 months, longer than 12 months, or it did not revert. In some embodiments, the subject is experienced hair loss after pregnancy, and if so, the survey asks if the hair loss stopped within a month, 1 to 3 months, 3 to 6 months, 6 to 12 months, longer than 12 months, or it did not stop. In some embodiments, the subject experienced stretch marks during pregnancy, and if so, the survey asks how long it took the stretch marks to go away, including without limitations, within a month, 1 to 3 months, 3 to 6 months, 6 to 12 months, longer than 12 months, or they did not go away.

[0296] In some embodiments, the survey asks how long after giving birth did it take for the subject’s period to return, including without limitations, within a month, 1 to 3 months, 3 to 6 months, 6 to 12 months, longer than 12 months, or other. In some embodiments, the subject’s period may be different following childbirth.

[0297] In some embodiments, a subject is experiencing or at risk for infertility. Infertility comprises an inability to become pregnant (conceive) after one year of trying to conceive. In some embodiments, subjects experiencing or at risk for infertility is experiencing HMB or AUB, is at risk for HMB or AUB, and / or has or be at risk for a menstrual cycle disorder. In some embodiments, the subject is tried to get pregnant, tried to get pregnant for more than 6 months in a row without succeeding, sought treatment for fertility in any clinic, had a fertility work up, been diagnosed with infertility, or had a partner diagnosed with infertility.

[0298] In some embodiments, a subject has a family history of fertility or infertility. In some embodiments, the mother of the subject’s age when they had their first child is at least 13, at least 15, at least 20, at least 25, at least 30, at least 35, or at least 40 years of age. In some embodiments, the mother of the subject’s age when they had their last child is at least 13, at least 15, at least 20, at least 25, at least 30, at least 35, or at least 40 years of age. Menopause and Perimenopause

[0299] In some embodiments of method provided herein, the digital biomarker comprises a phenotypical or behavioral property related to menopause or perimenopause. In some embodiments, a subject is experiencing menopause. In some embodiments, a subject experiencing menopause has entered menopause early. Early menopause is menopause which begins before a threshold age, such as 40 years of age. Early menopause is caused by genetic factors, chemical factors (e.g., chemotherapy), a combination of genetic and chemical factors, or unknown factors. In some embodiments, a subject has entered menopause at 25 years of age, 26 years of age, 27 years of age, 28 years of age, 29 years of age, 30 years of age, 31 years of age, 2020373103   25 Mar 2026 32 years of age, 33 years of age, 34 years of age, 35 years of age, 36 years of age, 37 years of age, 38 years of age, 39 years of age, 40 years of age, 41 years of age, 42 years of age, 43 years of age, 44 years of age, 45 years of age, 46 years of age, 47 years of age, 48 years of age, 49 years of age, 50 years of age, or over 50 years of age.

[0300] In some embodiments, the survey collects information on menopause and perimenopause. In some embodiments, the survey collects information on Premature Ovarian Failure, use of hormone tests, changes to menstrual cycle, vasomotor symptoms, sleep issues, mood changes, physical symptoms, genitourinary symptoms, appearance changes, life changes due to menopause, hormone replacement therapy, bone loss, osteoporosis, or treatments for osteoporosis. In some embodiments, the hormone test is for Follicle-stimulating hormone (FSH), Estrogen (estradiol), Thyroid-stimulating hormone (TSH), or Anti-Mullerian hormone (AMH). In some embodiments, the treatments for osteoporosis comprise Vaginal estrogen, Low-dose antidepressants, Gabapentin (Neurontin, Gralise, others), Clonidine (Catapres, Kapvay, others), or other medications to prevent or treat osteoporosis. In some embodiments, the survey asks the subject if the hormone tests indicated any of the following: currently in menopause, not in menopause, or not sure (test results were unclear).

[0301] In some embodiments, the subject recently noticed changes that the subject think might indicate movement towards menopause, with the options being yes, no, or not sure. In some embodiments, these changes occurred within 1, ,2 ,3 ,4 ,5, 6, or 7 years.

[0302] In some embodiments, the subject’s menstrual cycle recently become less regular than it used to be. In some embodiments, the cycle changed such that the subject loses more blood, loses less blood, the blood loss is irregular, there are more days in between periods, there are less days in between periods, the amount of flow has changed, the consistency of flow has changed.

[0303] In some embodiments, the subject experienced any of the following vasomotor symptoms (due to constriction or dilation of blood vessels): hot flashes, night sweats, feeling flushed, racing heart, perspiration, chills, or heart palpitations. In some embodiments, the subject experienced sleep issues such as insomnia, waking up during the night, sweating in the night, difficulty falling asleep due to physical discomfort, or daytime drowsiness. In some embodiments, the subject experienced mood changes such as irritability, fatigue, confusion, anger, tension, depression, anxiety, mood swings, or changes in libido (sex drive). In some embodiments, the subject experienced physical symptoms such as headaches, sore / tender breasts, muscle aches and pains, joint pain, or digestive issues. In some embodiments, the 2020373103   25 Mar 2026 subject experienced genitourinary (vaginal & urethra) symptoms such as vaginal dryness, vaginal itchiness, vaginal discharge, burning with urination, frequent urination, recurrent UTIs, urinary incontinence, pain with intercourse, or shortening and / or tightening of the vaginal canal. In some embodiments, the subject experienced changes to appearance such as undesired weight gain, changes to the subject’s silhouette (i.e. wider waist), appearance of acne, thinner hair on the top of the subject’s head, thicker facial / body hair, less-even skin tone, or loss of skin elasticity.

[0304] In some embodiments, the subject made any of the following life changes in anticipation of the transition to menopause, or to counteract the apparent effects of menopause: increased calcium intake, increased vitamin D intake, quit smoking, quit drinking alcohol, reduced alcohol intake, increased physical activity, adopted dietary changes aimed toward weight loss, increased my performance of high impact weight-bearing exercises (i.e. running, skipping, dancing), started to take non-hormone medications, or started to use topic hormone therapies (normally an estrogen cream / insert / gel that is applied to the vagina).

[0305] In some embodiments, the subject sought hormone replacement therapy (HRT) in association with menopause. HRT may have started within 1 year, 2, years, 3, years, 4, years, 5, years, 6, years, 7 years, 8 years, 9 years or 10 years. In some embodiments, the subject experienced adverse outcomes after completing HRT such as bloating / swelling / tenderness in breasts, headaches and / or migraines, nausea, indigestion, unexpected vaginal bleeding, mood swings, acne, breast cancer, ovarian cancer, uterine cancer, blood clots, heart disease, or stroke.

[0306] In some embodiments, the subject experienced bone loss. In some embodiments, the subject is (or suspect to have) osteoporosis. In some embodiments, the subject has been diagnosed with osteoporosis by a doctor. In some embodiments, the subject used any of the following treatments: vaginal estrogen, low-dose antidepressants, Gabapentin (Neurontin, Gralise, others), Clonidine (Catapres, Kapvay, others), or medications to prevent or treat osteoporosis. In some embodiments, the subject used alternative treatments such as acupuncture, yoga, bioidentical hormones, plant estrogens (phytoestrogens), black cohosh, aromatherapy, meditation & mindfulness, hypnosis, vigorous exercise, cognitive behavioral therapy program, therapeutic massage, or isoflavens (soy). Medical History

[0307] In some embodiments of method provided herein, the digital biomarker comprises a phenotypical or behavioral property related to the medical history of the subject. In some embodiments, the survey collects information on medical history. In some embodiments, the 2020373103   25 Mar 2026 medical history is the age of first period or the age of mother’s first period. In some embodiments, the information is a diagnosis of abnormal menstrual bleeding, abnormal pap, anemia, asthma, premature menarche, delayed menarche, hypothalamic amenorrhea, dysmenorrhea, eating disorder, endometriosis, polycystic ovarian syndrome, premature menopause, ovarian failure, ovarian cysts, fibroids, genital prolapse, genital warts, hepatitis, HIV / AIDS, STIs, migraine headaches, obesity, substance abuse, cervical cancer, breast cancer, fibrocystic breast disease, dense breasts, ovarian cancer, endometrial cancer, pelvic inflammatory disease, infertility, diminished ovarian reserve, chronic or frequent UTIs, ectopic pregnancy, heart disease, Type I Diabetes, Type II Diabetes, hypothyroidism, hyperthyroidism, Hashimoto’s, autoimmune conditions (lupus, MS, rheumatoid arthritis), or a combination thereof. In some embodiments, the information is a family history of clotting disorders, factor v leiden, hemophilia, endometriosis / adenomyosis, ovarian cysts, polycystic ovarian syndrome, fibroids, breast cancer, ovarian cancer, endometrial cancer, pelvic inflammatory disease, infertility, diminished ovarian reserve, chronic or frequent UTIs, ectopic pregnancy, preeclampsia, heart disease, Type I Diabetes, Type II Diabetes, or autoimmune conditions (Lupus, MS, rheumatoid arthritis). In some embodiments, the survey collects information on a past surgery. In some embodiments, the past surgery is an abdominal surgery, appendectomy, bladder suspension, breast biopsy, breast lumpectomy, cervical conization, cervical cryosurgery, laser treatment, cervix, cesarean section (C-section), cholecystectomy, colposcopy, diagnostic laparoscopy, dilation and curettage, endometrial ablation, hemorrhoidectomy, hernia repair, hysterectomy, hysteroscopy, myomectomy, oophorectomy, ovarian cystectomy, prolapse surgery, therapeutic laparoscopy, LEEP procedure (Loop electrosurgical excision), low back pain surgery, mastectomy, partial colectomy, releasing of peritoneal adhesions (scar tissue removal), tonsillectomy, tubal ligation, or a combination thereof. In some embodiments, the medical history is a test for BRCA1 / BRCA2. If the subject has been tested for BRCA1 / BRCA2 gene mutations, the survey asks which provider the subject used, including without limitations, Myriad Genetics, Color Genomics, Invitae, 23andme, Don’t Know, or other provider. In some embodiments, the subject is BRCA1 / BRCA2 positive (found to have mutations linked to breast and ovarian cancer in the subject’s BRCA genes). In some embodiments, a subject takes medications, including over-the-counter medications or prescription medications. In some embodiments, a medication is paracetamol, acetaminophen, aspirin, ibuprofen, a COX-2 inhibitor (e.g., celecoxib or rofecoxib), an anti-inflammatory drug (e.g., naproxen mefanamix acid, aleve, 102anax102102102, 102anax102102, ketoprofen, anaprox), a narcotic drug (e.g., 2020373103   25 Mar 2026 hydrocodone, codeine, morphine, oxycontin, hydrocodone, Demerol, or meperidine), another pain-killing drug such as one aimed at the nerves or central nervous system (e.g., amitriptyline, nortriptyline, gabapentin, pregabalin, or lamotrigine), a muscle relaxant (e.g., diazepam, temazepam, soma, lorzone, fexmid, or buscopan), an anti-anxiety or anti-depressant drug (e.g., citalopram, escitalopram, fluoxetine, fluvoxamine, paroxetine, diazepam, 103anax103103, 103anax103, luvox, paxil, 103anax103, 103anax, valium, amitriptyline, nortriptyline, SSRIs including sertraline), a stimulant (e.g., Adderall, Ritalin, Vyanese or methylphenidate), an HPV vaccine (e.g., Gardasil), estrogen-blockers, an herbal medicine or supplement, a vitamin or mineral supplement, or a multivitamin supplement. In some embodiments, a subject is currently taking one or more medications. In some embodiments, a subject has taken one or more medications within the past 1, 2, 3, 4, 5, or 6 months.

[0308] In some embodiments, the survey collects information about access to health care. In some embodiments, the survey collects information about health insurance, complementary and alternative medicine, costs associated with seeking health care, access to a vehicle, healthcare providers, most recent healthcare visit, preventative care, access to a medical professional.

[0309] In some embodiments, the complementary and alternative medicine is acupuncture, Ayurveda, homeopathy, naturopathy, traditional Chinese medicine, chiropractic and osteopathic medicine, dietary supplements, energy therapies (e.g. reiki), meditation, or body movement therapies (e.g. yoga, tai chi).

[0310] In some embodiments, the preventative care is a flu shot, a mammogram, a Pap smear, an HPV test, or a physical exam. In some embodiments, the subject receives preventative care at least about every 1, 2, ,3 ,4, 5, 6, 7, 8, ,9 or 10 years. In some embodiments, the subject needs to schedule an appointment for preventative care at least about 1 week, 2 weeks, 3 weeks, 4, weeks, 1 month, 2 months, 3, mores, 5 moths, 5 months, 6 months, or more than 6 months in advance of an appointment.

[0311] In some embodiments, the medical professional is a primary care physician or an OBGYN. In some embodiments, the subject is a Primary Care Physician to call when the subjects is sick or has a question about health. In some embodiments, the subject is an OBGYN (Obstetrician / Gynecologist) for the subject’s reproductive needs. In some embodiments, the Primary Care Physician or OBGYN is located a distance away from the subject, including without limitations, less than 5 miles away, 5-10 miles away, 11-20 miles away, more than 20 miles away, more than 50 miles away, or more than 100 miles away. In some embodiments, the subject has a copay for an appointment. In some embodiments, the subject is regular access 2020373103   25 Mar 2026 (owns or leases) to a vehicle. In some embodiments, the survey asks how far away, in miles, is the closest hospital to the subject. In some embodiments, the subject is normally travel to a doctor’s appointment, including without limitations, by walking, public transportation, bike, drive, ask a family member / friend with a car to take me, shared ride service, or other. In some embodiments, the friend or family member joins the subject during doctor’s visits.

[0312] In some embodiments, the subject is a budget in mind for annual healthcare costs and if so, the survey asks about the subject’s budget. In some embodiments, the survey asks which of the following providers the subject is most likely to interact with during a medical appointment: nurse, PA (Physician’s Assistant), MA (Medical Assistant), or Doctor (MD). In some embodiments, the survey asks a subject how easy it is to get a referral from the subject’s primary care doctor for a specialist.

[0313] In some embodiments, the survey asks when the last time a subject went to the ER (Emergency Room). In some embodiments, the survey asks how many times a subject has been to the ER. In some embodiments, the survey asks when the last time a subject went to Urgent Care. In some embodiments, the survey asks how many times a subject has been to Urgent Care.

[0314] In some embodiments, the survey asks what the subject would be most likely to do if the subject had some abnormal symptoms related to reproductive health, including without limitations, call a primary care doctor’s office to schedule an appointment; call a gynecologist’s office to schedule an appointment; find the nearest Planned Parenthood for a consult; find the nearest minute clinic for a consult; wait to see if it gets worse, and if it does, find an urgent care clinic for a walk-in; wait to see if it gets worse, and if it does, find the closest hospital with an Emergency Room; seek on-line resources and wait to see how it develops; seek alternative at-home remedies to see if it resolves without escalating to a doctor; or other.

[0315] In some embodiments, the survey asks on a scale of 1 to 5, 1 being no consideration, 5 being heavy consideration, when the subject thinks about seeking medical care, how much consideration does the subject give to the following factors: co-pay, out of pocket costs before deductible, uncertainty around cost of tests prescribed, difficulty communicating with the doctor, wait time at doctor’s office, commute to doctor’s office, anxiety about results, costs of possible medications prescribed, having to take time off of work to go to a doctor, finding childcare to go to a doctor, or other factors.

[0316] In some embodiments, the survey asks the subject which plan best describes the subject’s health insurance status, including without limitations, Health maintenance organization; Preferred provider organization; Point of service plan; Exclusive provider organization plan; 2020373103   25 Mar 2026 High deductible health plan; Health Savings Account; Individual health plan purchased on the health insurance marketplace;; VA care; Medicaid; Medicare; COBRA; or a combination thereof.

[0317] In some embodiments, a subject has a strong pelvic floor or a weak pelvic floor. Strength of pelvic floor muscles is determined by placing one or more fingers just above the tailbone and above the intergluteal cleft, squeezing the pelvic floor muscles, and feeling the strength of the movement felt.

[0318] In some embodiments, a subject has control of their bladder, sometimes loses control of their bladder, or regularly lose control of their bladder. In some embodiments, a subject experiences leaking with physical activity such as exercise, bending, sneezing, or coughing. In some embodiments, a subject experiences a sudden, intense urge to urinate, which present with a loss of control of the bladder or an uncontrollable flow of urine. In some embodiments, a subject experiences a frequent or constant need to urinate, which present without the ability to completely empty the bladder. In some embodiments, a subject experiences a slow leak of urine even when they believe their bladder has been emptied. In some embodiments, the survey asks if a physical therapist or doctor has ever confirmed the strength of the subject’s pelvic floor. In some embodiments, the subject respond in the following ways: strong; weak; somewhere-in-between; or no. In some embodiments, the survey asks the subject to describe subject’s pelvic floor strength as strong, weak, somewhere-in-between, or don’t know, using FIG. 20 as a reference.

[0319] In some embodiments, a subject has had a vaginal yeast, viral, or bacterial infection in the past. In some embodiments, a subject has had a vaginal yeast, viral, or bacterial infection i...

Claims

1. A method of producing a desired preparation for assessment of menstrual health,comprising:(a) receiving a fluid sample collected from the vaginal cavity of a subject, the fluid sample comprising one or more components;(b) contacting the fluid sample, with a buffer solution under conditions to preserve one or more components of the fluid sample;(c) determining an expression level of a sample component from the one or more components of the fluid sample;(d)     obtaining survey data from the first subject and from a group of subjects,wherein the survey data comprises one or more features selected from the group consisting of a menstrual cycle phenotype, a physical body characteristic, a disease or condition, a medical treatment, demographic information, lifestyle information, ancestry, sexuality, menstrual management, health care usage, health care access, and health record data relating to said group of subjects; and(e) stratifying the group of subjects into subgroups based on said one or more features;(f) assigning said first subject to one of said subgroups based on said one or more features;(g) comparing said expression level of step (c) to a reference expression level of the sample component to generate a comparison, wherein the reference expression level of the sample component is obtained from the subgroup to which said first subject was assigned; and(h)    determining an assessment of menstrual health using the comparison obtained instep (g).

2. The method of claim 1, wherein the survey data comprises at least one of one or morefeatures, wherein at least one of the one or more features comprises a factor selected from the list consisting of age, location of residence, natural hair color, annual income, level of education, relationship status, frequency of consensual physical touch (outside of sexual contact), current living situation, religious background, current religious affiliation, military service, mental health, menstruation phenotype, pregnancy state, number of pregnancies, number and type of tampons or other menstrual products used during period cycle, height, weight, relationship2020373103   17 Jun 2026status, birth sex, race, ethnicity, sexual orientation, gender, hormone therapy, alcohol consumption, cigarette consumption, marijuana consumption, hemp / CBD consumption, exercise, diet, stress level, sleep, fertility treatments, infertility, preeclampsia, pre-term labor, miscarriage, abortion, vaginal birth, c-section, live births, mother’s pregnancy history, age of first menstruation, mother’s age of first menstruation, polycystic ovarian syndrome, ovarian cysts, fibroids, breast cancer, ovarian cancer, endometrial cancer, pelvic inflammatory disease, chronic or frequent UTIs, ectopic pregnancy, heart disease, disability, type I diabetes, type II diabetes, autoimmune conditions (lupus, MS, rheumatoid arthritis), diminished ovarian reserve, endometriosis / adenomyosis, family disease history, position of uterus, painful periods, heavy bleeding during period, irregular bleeding on or off period, short interval between periods, chronic pelvic pain, lower abdominal or back pain, pain during penetrative sex, pain while defecating, often with cycles of diarrhea and constipation, bloating, nausea, and vomiting, groin pain, pain during exercise, vaginal yeast, viral, or bacterial infection, vaginal infection treated with antibiotics, medication usage, freeze preservation of eggs, strength of pelvic floor, loss of bladder control, birth control usage and type, duration of hormonal birth control usage, sexual history, likelihood of orgasm during sex, medication or supplement or device usage for sexual enhancement, frequency of sex, occurrence of sexually transmitted infection, abnormal pap smear, age of first sexual intercourse, number of sexual partners, sex during period, usage of emergency contraception, current menstrual bleeding pattern, typical menstrual bleeding pattern, cycle length, number of days of bleeding, consistency of bleeding pattern over menstrual cycles, ovulation cycle, pain during menstrual window, type and level of pain experienced during menstrual window, pain outside menstrual window, type and level of pain experienced outside menstrual window, diet interaction with menstrual-associated pain, activity or treatment used for pain relief for menstrual-associated pain, spasms in uterus not during menstrual window, onesided, low belly pain during ovulation (mittelschmerz), rectal pain during period or ovulation, number of sanitary products used together for menstrual bleeding, passing blood clots during menstruation, menstrual bleeding level during sleep, menstrual bleeding level during day, interference of menstruation with typical daily activities, days of heavy flow during menstruation, pattern of bleeding during heavy flow days, pattern of bleeding during lighter flow days, pattern of bleeding throughout menstrual window, waist size, hip size, bust size, weight gain potential, weight loss potential, muscle mass, potential to increase or decrease muscle mass, areas of fat gain or accumulation, body areas having coarse hair, color of menstrual fluid, consistency of menstrual fluid, odor of menstrual fluid, feel of menstrual fluid, hair loss, acne,2020373103   17 Jun 2026body hair, history of environmental exposure, family history of disease, use of antibiotics, sex education, attitudes toward menstruation, health insurance, ER visits, and preventative care practices.

3. The method of any one of claims 1 or 2, wherein the reference level is obtained from ahealthy control subject or an average level from a group of healthy control subjects.

4. The method of any one of claims 1 to 3, wherein the one or more components of the fluidsample preserved are selected from the group consisting of intact cells from the vaginal-cervical space, protein, metabolite, DNA, RNA or any combination thereof.

5. The method of any one of claims 1 to 4, wherein step (c) comprises RNA sequencing.

6. The method of any one of claims 1 to 5, further comprising repeating steps (a)-(e) fortwo or more longitudinal samples from the subject.

7. The method of any one of claims 1 to 6, wherein the fluid sample is received in a samplecollector selected from the group consisting of a pad, a tampon, a vaginal cup, a cervical cap, a menstrual disk, a cervical disk, a sponge, and an interlabial pad.

8. The method of any one of claims 1 to 7, wherein the fluid sample is obtained from thesubject during the subject's menstrual window.

9. The method of any one of claims 1 to 8, wherein the assessment of menstrual healthindicates that the subject has a menstrual cycle disorder.

10. The method of claim 9, wherein the menstrual cycle disorder is heavy menstrual bleeding (HMB).

11. A method of producing a desired preparation for assessment of menstrual health, comprising:(a) receiving a fluid sample collected from the vaginal cavity of a subject, the fluid sample comprising one or more types of cells;2020373103   17 Jun 2026(b) contacting the fluid sample with a buffer solution under conditions suitable to maintain one or more cell types in a substantially intact state;(c) separating one cell type in the fluid sample from the remaining fluid sample;(d)    determining the level or presence of one or more components selected from thegroup consisting of nucleic acids, proteins and metabolites; and(e) comparing the presence or level obtained in step (d) with a reference to provide the assessment of menstrual health, wherein the reference is at least partially selected based on survey results12. The method of claim 11, wherein step (d) comprises determining an expression level of one or more markers in the one or more components in the one cell type and step (e) comprises applying a classifier algorithm to the expression level of one or more markers to calculate a metric that quantifies the difference between the expression level and a reference expression level.

13. The method of claim 11 or claim 12, wherein the one or more cell types are selected from the group consisting of leukocytes, erythrocytes, endothelial cells, epithelial cells, stromal cells, stem cells, and any combinations thereof.

14. The method of claim 11, wherein the survey results comprise information on one of more digital biomarker selected from the group consisting of a menstrual cycle phenotype, a physical body characteristic, a disease or condition, a medical treatment, demographic information, lifestyle information, ancestry, sexuality, menstrual management, health care usage, health care access, and health record data relating to the subject.

15. The method of claim 14, wherein at least one of the one or more digital biomarker comprises a factor selected from the list consisting of age, location of residence, natural hair color, annual income, level of education, relationship status, frequency of consensual physical touch (outside of sexual contact), current living situation, religious background, current religious affiliation, military service, mental health, menstruation phenotype, pregnancy state, number of pregnancies, number and type of tampons or other menstrual products used during period cycle, height, weight, relationship status, birth sex, race, ethnicity, sexual orientation, gender, hormone therapy, alcohol consumption, cigarette consumption, marijuana consumption, hemp / CBD2020373103   17 Jun 2026consumption, exercise, diet, stress level, sleep, fertility treatments, infertility, preeclampsia, preterm labor, miscarriage, abortion, vaginal birth, c-section, live births, mother’s pregnancy history, age of first menstruation, mother’s age of first menstruation, polycystic ovarian syndrome, ovarian cysts, fibroids, breast cancer, ovarian cancer, endometrial cancer, pelvic inflammatory disease, chronic or frequent UTIs, ectopic pregnancy, heart disease, disability, type I diabetes, type II diabetes, autoimmune conditions (lupus, MS, rheumatoid arthritis), diminished ovarian reserve, endometriosis / adenomyosis, family disease history, position of uterus, painful periods, heavy bleeding during period, irregular bleeding on or off period, short interval between periods, chronic pelvic pain, lower abdominal or back pain, pain during penetrative sex, pain while defecating, often with cycles of diarrhea and constipation, bloating, nausea, and vomiting, groin pain, pain during exercise, vaginal yeast, viral, or bacterial infection, vaginal infection treated with antibiotics, medication usage, freeze preservation of eggs, strength of pelvic floor, loss of bladder control, birth control usage and type, duration of hormonal birth control usage, sexual history, likelihood of orgasm during sex, medication or supplement or device usage for sexual enhancement, frequency of sex, occurrence of sexually transmitted infection, abnormal pap smear, age of first sexual intercourse, number of sexual partners, sex during period, usage of emergency contraception, current menstrual bleeding pattern, typical menstrual bleeding pattern, cycle length, number of days of bleeding, consistency of bleeding pattern over menstrual cycles, ovulation cycle, pain during menstrual window, type and level of pain experienced during menstrual window, pain outside menstrual window, type and level of pain experienced outside menstrual window, diet interaction with menstrual-associated pain, activity or treatment used for pain relief for menstrual-associated pain, spasms in uterus not during menstrual window, one-sided, low belly pain during ovulation (mittelschmerz), rectal pain during period or ovulation, number of sanitary products used together for menstrual bleeding, passing blood clots during menstruation, menstrual bleeding level during sleep, menstrual bleeding level during day, interference of menstruation with typical daily activities, days of heavy flow during menstruation, pattern of bleeding during heavy flow days, pattern of bleeding during lighter flow days, pattern of bleeding throughout menstrual window, waist size, hip size, bust size, weight gain potential, weight loss potential, muscle mass, potential to increase or decrease muscle mass, areas of fat gain or accumulation, body areas having coarse hair, color of menstrual fluid, consistency of menstrual fluid, odor of menstrual fluid, feel of menstrual fluid, hair loss, acne, body hair, history of environmental exposure,2020373103   17 Jun 2026family history of disease, use of antibiotics, sex education, attitudes toward menstruation, health insurance, ER visits, and preventative care practices.

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