Biomarker for adolescent depressive disorder diagnosis and application

Abstract

The present invention provides a method for the specific diagnosis of a teenager male suffering from a major depressive disorder in a teenager male population, comprising determining the expression level of a biomarker in a sample from a teenager male subject and comparing to a reference value, wherein the teenager male subject is suspected to suffer from the major depressive disorder, the biomarker comprises HER2, and reduction of the expression level of the biomarker compared with a reference value prompts that the teenager male subject suffers from the major depressive disorder. The invention also provides a method, a composition, a test strip, a test paper card and / or a kit for distinguishing or differentially diagnosing whether a teenager male subject suffers from major depressive disorder or not by utilizing the detection of the biomarker. The method composition, the test strip, the test paper card and / or the kit can specifically diagnose the major depressive disorder, and have excellent accuracy and sensitivity.

Description

Technical Field

[0001] The present invention relates to the field of medical diagnosis, specifically to biomarkers for diagnosing major depressive disorder in adolescents, more specifically to biomarkers for specifically diagnosing major depressive disorder in adolescent males, and biomarkers for differentiating and differentially diagnosing adolescent males with major depressive disorder from other mental illnesses such as bipolar disorder and schizophrenia. The present invention also relates to methods, compositions, test strips, test cards, and / or kits for diagnosing adolescent males with major depressive disorder by detecting such biomarkers. Background Art

[0002] Mental illness, also known as mental disorder, is a general term for a class of all pathological mental activities, which is manifested as a syndrome characterized by clinically significant individual cognitive, emotional regulation, or behavioral disorders. Common mental illnesses include Neurodevelopmental Disorders (ND), Depressive Disorders (DD), Schizophrenia (SZ), Anxiety Disorders (AD), Bipolar Depressive Disorders (BDD), and other conditions.

[0003] Currently, the classification criteria for mental illnesses include the DSM system based on the American Psychiatric Association, the ICD system based on the World Health Organization, the CCMD system based on China, etc. These classification systems have certain differences, and these systems are all based on clinical symptom assessment and lack objective biological diagnostic indicators. That is, subjects with a group of similar symptoms are classified into the category of the same disease, and this group of similar symptoms may be caused by completely different biological factors / pathological factors. On the contrary, although subjects with different symptoms are classified as different diseases based on clinical symptom assessment, these symptoms may be caused by the same biological factors / pathological factors. This brings difficulties to the accurate treatment of mental illnesses.

[0004] Among various mental illnesses, depressive disorder is a complex neuropsychiatric disorder. According to the latest DSM-V criteria, depressive disorder can be further divided into major depressive disorder (MDD, typical depression), persistent depressive disorder (dysthymia), premenstrual dysphoric disorder, etc. Major depressive disorder is the most common type of depressive disorder. Patients with major depressive disorder may have the same or similar manifestations as other mental illnesses such as bipolar disorder and schizophrenia in some cases. In particular, major depressive disorder shows an increasing trend year by year among adolescents aged 12 - 18. Major depressive disorder in adolescents is usually not detected in time or misdiagnosed as "adolescent rebellion" or "mood swings", thus delaying treatment. Currently, the commonly used diagnostic criteria for major depressive disorder in adolescents in clinical practice include psychological assessment, scale tests (such as PHQ-9, CDI), doctor interviews, and clinical observations, and these methods rely more on the doctor's experience and judgment.

[0005] Therefore, there is an urgent need for biomarkers for the diagnosis of major depressive disorder, especially biomarkers that can be used clinically to distinguish or differentially diagnose major depressive disorder from other mental illnesses with similar clinical manifestations to major depressive disorder. Finding and establishing one or a set of clear biomarkers to achieve the objective diagnosis of major depressive disorder is of great clinical significance. Summary of the Invention

[0006] Based on mass spectrometry analysis and ELISA verification, the present invention analyzes the differentially expressed proteins in samples from adolescent patients with major depressive disorder to find biomarkers for the diagnosis of adolescent major depressive disorder. The present invention unexpectedly finds that the expression of HER2 protein has gender differences, and it is differentially expressed in adolescent male patients with major depressive disorder. Moreover, the expression of HER2 protein in adolescent males with major depressive disorder is statistically significantly different from its expression in healthy adolescent males, adolescent males with bipolar disorder, and adolescent males with schizophrenia. The present invention provides an objective diagnostic basis for specifically diagnosing, differentiating, and differentially diagnosing adolescent males with major depressive disorder clinically, and for differentiating and differentially diagnosing adolescent males with major depressive disorder from healthy adolescent males, adolescent males with bipolar disorder, and adolescent males with schizophrenia clinically.

[0007] The present invention provides the use of a reagent for detecting HER2 in a sample from an adolescent male subject in the preparation of a kit, test strip, or test card for specifically diagnosing, differentiating, and / or differentially diagnosing adolescent males with major depressive disorder in the adolescent male population.

[0008] The present invention further provides a composition, test strip, test card and / or kit for specific diagnosis, differentiation and / or differential diagnosis of adolescent males suffering from major depressive disorder, which are prepared based on a detection reagent for HER2 protein.

[0009] The method, composition, test strip, test card and / or kit of the present invention can specifically diagnose whether an adolescent male has a depressive disorder in the adolescent male population, and in particular, can distinguish and differentially diagnose adolescent males suffering from major depressive disorder from other mental diseases such as bipolar disorder and schizophrenia. The method, composition, test strip, test card and / or kit of the present invention have excellent diagnostic specificity, sensitivity and accuracy for whether an adolescent male has a depressive disorder. Description of the Drawings

[0010] Figure 1 : Mass spectrometry signal intensity of HER2 in healthy groups (HC) and adolescent males and females with major depressive disorder (MDD).

[0011] Figure 2 : Mass spectrometry signal intensity of HER2 in healthy groups (HC) and adolescent males and females with schizophrenia (SZ).

[0012] Figure 3 : Mass spectrometry signal intensity of HER2 in healthy groups (HC), adolescent males with major depressive disorder (MDD), schizophrenia (SZ) and bipolar disorder (BD).

[0013] Figure 4 : ELISA detection of the content of HER2 in the blood of healthy groups (HC) and adolescents with major depressive disorder (MDD).

[0014] Figure 5 : ELISA detection of the content of HER2 in the blood of healthy groups (HC) and adolescent males and females with major depressive disorder (MDD).

[0015] Figure 6 : T-test chart of test results of healthy people and samples with major depressive disorder.

[0016] Figure 7 : ROC curve of test results of healthy people and samples with major depressive disorder. Detailed Description of the Invention

[0017] A. Definitions

[0018] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. References to techniques used herein are intended to refer to techniques as commonly understood in the art, including variations of those techniques or substitutions of equivalent techniques that are obvious to one of ordinary skill in the art. Although the following terms are believed to be well understood by one of ordinary skill in the art, the following definitions are set forth to better explain the present invention.

[0019] As used herein, the term "biomarker", also referred to as "biological marker", refers to a measurable indicator that reflects the biological state of a subject. Such a biomarker can be any substance (e.g., a gene or a protein) in a subject, provided that they are related to a specific biological state (e.g., a disease) of the subject being tested. A biomarker can be a single marker or a set of markers consisting of two or more biomarkers. The biomarker in the present invention refers to a protein.

[0020] As used herein, the names of the biomarkers for specifically diagnosing adolescent males with major depressive disorder are referred to the definitions in the UniProt protein database. In one embodiment, the following proteins all refer to proteins in the species Homo sapiens (Human).

[0021] The term "HER2" is a tyrosine kinase-type cell surface receptor, which is also referred to as receptor tyrosine-protein kinase erbB-2. The accession number of HER2 in the UniProt protein database is P04626. See specifically https: / / www.uniprot.org / uniprotkb / P04626 / entry. The terms "HER2", "HER2 protein" and "the biomarker of the present invention" can be used interchangeably herein.

[0022] HER2 is known to be a protein related to cancer. Only a few studies have mentioned the relationship between ErbB2 / HER2 and mental disorders. For example, in 2017, Mostaid MS et al. (Mostaid MS, Lee TT, Chana G, Sundram S, ShannonWeickert C, Pantelis C, Everall I, Bousman C. Peripheral Transcription of NRG-ErbB Pathway Genes Are Upregulated in Treatment-Resistant Schizophrenia. Front Psychiatry. 2017 Nov 6;8:225) reported that in the blood of patients with treatment-resistant schizophrenia, the transcriptome expression of HER2 was negatively correlated with the disease progression; however, there was no difference in the expression abundance of HER2 between the healthy group and the patient group. In 2018, Wang XD et al. (Wang XD, Su YA, Guo CM, Yang Y, Si TM. Chronic antipsychotic drug administration alters the expression of neuregulin 1beta, ErbB2, ErbB3, and ErbB4 in the rat prefrontal cortex and hippocampus. Int J Neuropsychopharmacol. 2008 Jun;11(4):553-61) reported that the antipsychotic drugs haloperidol and clozapine could promote the increase of ErbB2 protein in the prefrontal cortex of rats. In 2016, Dang, R. et al. (Dang, R., Cai, H., Zhang, L., Liang, D., Lv, C., Guo, Y., et al. (2016). Dysregulation of Neuregulin-1 / ErbB signaling in the prefrontal cortex and hippocampus of rats exposed to chronic unpredictable mild stress. Physiology & Behavior, 154, 145–150) found that when rats entered a depressive-like state, the expressions of ErbB2 and pErbB2 were both inhibited, and on the contrary, the expression of pErbB4 increased. Treatment with the antidepressant sertraline enhanced the NRG1 / ErbB signaling pathway and partially reversed the stress-induced behavioral changes and the disorder of the NRG1 / ErbB system in rats.The relationship between HER2 protein and major depressive disorder, especially in adolescent males with major depressive disorder, has not been reported to date.

[0023] In this article, the diagnostic criteria for the diseases mentioned are based on the Diagnostic and Statistical Manual of Mental Disorders, 5th Edition (DSM-V) of the American Psychiatric Association. According to the DSM-V diagnostic criteria, depressive disorders are an independent category of diseases and can be further classified into 8 subtypes, including Disruptive Mood Dysregulation Disorder, Major Depressive Disorders (MDD, typical depression), Persistent Depressive Disorder (dysthymia), Premenstrual Dysphoric Disorder, etc. In one embodiment, the clinical classification of MDD is based on the DSM-V diagnostic criteria.

[0024] In this article, the terms "differentially expressed proteins", "differentially expressed protein" or "differentially expressed protein" (DEPs) refer to proteins with a fold change in expression > 2 or < 0.5 and p < 0.05 in a two-tailed t-test after analyzing plasma samples. For example, if comparing the protein expression changes in MDD subjects relative to healthy controls, proteins with a fold change in expression > 2 or < 0.5 (p < 0.05 in a two-tailed t-test; data is normalized) in MDD subjects are considered to be differentially expressed proteins.

[0025] In this article, the terms "diagnose", "diagnosed" or "diagnosing" refer to methods of detecting and / or identifying the case status of a subject and differentiating and / or determining whether a subject has a specific disease.

[0026] The term "specifically diagnosing adolescent males with major depressive disorder" refers to specifically diagnosing whether a specific adolescent male in the adolescent male population has major depressive disorder by detecting the biomarker HER2 discovered in the present invention. The term "specifically diagnosing adolescent males with major depressive disorder" includes specifically diagnosing adolescent males with major depressive disorder from the adolescent male population and excluding the possibility that a specific adolescent male in the adolescent male population has major depressive disorder.

[0027] The term "differentiating and / or differential diagnosing adolescent males with major depressive disorder from adolescent males with bipolar disorder and / or schizophrenia" refers to specifically diagnosing adolescent males with major depressive disorder from patients suspected of having major depressive disorder, but who also exhibit symptoms similar to those of major depressive disorder, such as patients with bipolar disorder and schizophrenia, by detecting the biomarker HER2 discovered by the present invention. The term "differentiating and / or differential diagnosing adolescent males with major depressive disorder from adolescent males with bipolar disorder and / or schizophrenia" includes specifically diagnosing adolescent males with major depressive disorder from patients suspected of having major depressive disorder, but who also exhibit symptoms similar to those of major depressive disorder, such as patients with bipolar disorder and schizophrenia; and excluding the possibility that a specific adolescent male has major depressive disorder from patients suspected of having major depressive disorder, but who also exhibit symptoms similar to those of major depressive disorder, such as patients with bipolar disorder and schizophrenia.

[0028] In this document, the terms "subject" and "patient" may be used interchangeably. Unless otherwise specified, a "subject" in this document refers to an adolescent between 10 and 18 years of age.

[0029] In this document, specific diagnosis and / or differentiation and / or differential diagnosis of major depressive disorder is carried out by comparing the expression level of the biomarker of the present invention with a reference value, and a decrease in the expression level of the biomarker compared with the reference value indicates that the adolescent male subject has major depressive disorder. In one embodiment, the reference value is the expression level of the biomarker in a sample from a healthy subject. Methods for measuring the expression level of the biomarker are known in the prior art.

[0030] In this text, "the expression level of the biomarker" can be determined by a variety of methods, including, for example, measuring protein concentration, measuring chemiluminescence value, etc. "The expression level of the biomarker is decreased compared with the reference value" can be expressed in a variety of forms. For example, the measured luminescence value is lower than the Cutoff value and the measured concentration is lower than the threshold concentration. The degree of decrease can be expressed as a multiple relationship, in the form of a percentage, or as a specific decreased value. In one embodiment, "the expression level of the biomarker is decreased compared with the reference value" means a decrease of 0.1 - 10 times, such as 0.1, 0.2, 0.3... 0.9, 1, 1.1, 1.2... 9.9 and 10 times. In one embodiment, "the expression level of the biomarker is decreased compared with the reference value" means a decrease of 5% - 90%, such as 5%, 6%, 7%... 89% and 90%. In one embodiment, "the expression level of the biomarker is decreased compared with the reference value" means that the measured protein concentration is lower than the set threshold concentration by values such as 1, 2, 3, 4... etc. In this text, the terms "reference value" and "set threshold" can be used interchangeably.

[0031] In this text, the term "sample" refers to a sample isolated from a subject's tissue or body fluid, including but not limited to, for example, whole blood or any blood component, plasma, serum, urine, cerebrospinal fluid, blood derivatives, blood cells, lymph fluid. Specifically, the sample can be in the form of a blood sample, plasma sample, or serum sample obtained from a subject suspected of having major depressive disorder.

[0032] In this text, the term "reagent capable of detecting the expression level of a biomarker" refers to a substance or means that can be used to determine whether the expression level of a biomarker in a sample obtained from a subject is increased, unchanged, or decreased. In this text, a reagent capable of detecting the expression level of a biomarker can be a binding partner that can target the biomarker and bind to the biomarker. For example, the binding partner can be an antibody or an antigen-binding fragment thereof that can target the biomarker and bind to the biomarker. The expression level of the biomarker is detected by a binding partner that targets the biomarker. For example, the expression level of the biomarker is detected by an antibody that can specifically bind to this biomarker. For another example, the expression level of the biomarker is determined by mass spectrometry, or an antibody-based immunoassay method, such as competitive immunoassay, non-competitive immunoassay, enzyme-linked immunosorbent assay (ELISA), immunohistochemical assay, chemiluminescence assay, Western blot assay, and Dot blot assay, etc.

[0033] The term "antibody" refers to an immunoglobulin molecule that contains at least one antigen recognition site and can specifically bind to an antigen. For example, an antibody can be a monoclonal antibody, a polyclonal antibody, an antibody fragment, a humanized antibody, a camel antibody, a chimeric antibody, etc. An antibody can be further modified to carry a detectable label, such as a label that can be detected by a chemiluminescence method.

[0034] In this document, unless the context clearly indicates otherwise, the singular forms "a", "an", and "the" include plural references.

[0035] In this document, the terms "optionally", "optional", or "optionally" mean that the subsequent described event or situation may or may not occur. For example, "Optionally, the test strip or test card further includes a detection part for detecting whether the biomarker binds to the reagent, and / or a part for quality control" means that the test strip or test card may further include a detection part for detecting whether the biomarker binds to the reagent, and / or a part for quality control; or this situation may not exist.

[0036] In this document, the terms "comprising", "including", "having", or "containing" and other variant forms thereof in this document are inclusive or open-ended and do not exclude other unlisted elements or method steps.

[0037] B. Compositions, test strips, test cards, and / or kits for specific diagnosis, differentiation, and / or differential diagnosis in adolescent males with major depressive disorder

[0038] The present invention has determined that HER2 can be used for specific diagnosis, differentiation, and / or differential diagnosis in adolescent males with major depressive disorder based on mass spectrometry and enzyme-linked immunosorbent assay.

[0039] The present invention performed data-independent acquisition (DIA) mass spectrometry analysis on samples including those from adolescent patients with major depressive disorder, adolescent patients with bipolar disorder, adolescent patients with schizophrenia, and healthy adolescents, and found that the mass spectrometry signal intensity of HER2 showed significant differences only between male and female adolescent patients with major depressive disorder and the healthy adolescent group. Through further verification by ELISA, it was found that there are gender differences in the expression of HER2, and there are significant differential expressions between adolescent male patients with major depressive disorder and healthy adolescent males. The repeatability, accuracy, and specificity of the detection results were further confirmed by chemiluminescence methods.

[0040] Accordingly, the present invention provides a composition, test strip, test card and / or kit for specific diagnosis, differentiation and / or differential diagnosis of adolescent males suffering from major depressive disorder, which is prepared based on a detection reagent for HER2 protein.

[0041] In a first aspect, the present invention further provides a composition, test strip, test card and / or kit for specific diagnosis, differentiation and / or differential diagnosis of adolescent males suffering from major depressive disorder, which is prepared based on a detection reagent for HER2 protein. Specifically,

[0042] In an exemplary aspect, the present invention provides a kit, test strip or test card for specifically diagnosing adolescent males suffering from major depressive disorder in the adolescent male population, comprising a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject suspected of suffering from major depressive disorder, wherein the biomarker comprises HER2, and a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject suffers from major depressive disorder, and the kit, test strip or test card further comprises instructions for use or an insert indicating that a decrease in the expression level of the biomarker compared to the reference value indicates that the adolescent male subject suffers from major depressive disorder.

[0043] In an exemplary aspect, the present invention provides a kit, test strip or test card for differentiating or differentially diagnosing adolescent males suffering from major depressive disorder and healthy adolescent males in the adolescent male population, comprising a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject suspected of suffering from major depressive disorder, wherein the biomarker comprises HER2, and a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject suffers from major depressive disorder, and the kit, test strip or test card further comprises instructions for use or an insert indicating that a decrease in the expression level of the biomarker compared to the reference value indicates that the adolescent male subject suffers from major depressive disorder.

[0044] In an exemplary aspect, the present invention provides a kit, test strip or test card for differentiating or differentially diagnosing adolescent males with major depressive disorder and adolescent males with schizophrenia in the adolescent male population, comprising a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder or schizophrenia, the biomarker includes HER2, and a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder. The kit, test strip or test card further includes instructions or inserts indicating that a decrease in the expression level of the biomarker compared to the reference value indicates that the adolescent male subject has major depressive disorder.

[0045] In an exemplary aspect, the present invention provides a kit, test strip or test card for differentiating or differentially diagnosing adolescent males with major depressive disorder and adolescent males with bipolar disorder in the adolescent male population, comprising a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder or bipolar disorder, the biomarker includes HER2, and a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder. The kit, test strip or test card further includes instructions or inserts indicating that a decrease in the expression level of the biomarker compared to the reference value indicates that the adolescent male subject has major depressive disorder.

[0046] In an exemplary aspect, the present invention provides a kit, test strip or test card for differentiating or differentially diagnosing adolescent males with major depressive disorder, adolescent males with bipolar disorder, and adolescent males with schizophrenia in the adolescent male population, comprising a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder, bipolar disorder or schizophrenia, the biomarker includes HER2, and a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder. The kit, test strip or test card further includes instructions or inserts indicating that a decrease in the expression level of the biomarker compared to the reference value indicates that the adolescent male subject has major depressive disorder.

[0047] According to the above aspects, the present invention further includes the following embodiments.

[0048] In one embodiment, the sample is selected from plasma, serum, urine, cerebrospinal fluid, blood derivatives, blood cells or lymph fluid. In one embodiment, the sample is a blood sample, preferably the sample is a plasma or serum sample. In one embodiment, the sample is a blood sample, plasma sample or serum sample obtained from a subject suspected of having or having a mental disorder. In one embodiment, the sample is a blood sample, plasma sample or serum sample obtained from a subject suspected of having or having major depressive disorder, schizophrenia or bipolar disorder.

[0049] In one embodiment, the age of the adolescent male population is 12 - 18 years old.

[0050] In one embodiment, a decrease in the expression level of HER2 by 0.1 - 10 times (such as 1 - 10 times, 2 - 9 times, 3 - 8 times, 4 - 7 times, 4 - 6 times, and 5 - 6 times) compared to the reference value indicates that the adolescent male subject has major depressive disorder. In one embodiment, a decrease in the expression level of HER2 by 5% - 90% (such as 10% - 90%, 30% - 80%, 40% - 70%, and 40% - 60%) compared to the reference value indicates that the adolescent male subject has major depressive disorder. In one embodiment, the expression level of HER2 is the concentration of HER2 protein in the sample. In one embodiment, a decrease in the concentration of HER2 protein by a value greater than 0.1 (such as 0.1, 0.2, 0.3, …… 1, 1.1, 1.2, …… 2, …… 99.8, 99.9, and 100, etc.) compared to the reference value indicates that the adolescent male subject has major depressive disorder.

[0051] In one embodiment, the reference value is the expression level of the biomarker in a sample from a healthy subject or from a standard sample. In one embodiment, the healthy subject may optionally be a healthy adolescent male subject, and / or the standard sample may optionally be a sample from a healthy adolescent male subject.

[0052] In the case where the biomarker to be detected is known, the method of preparing an antibody against this biomarker and further preparing a test strip and a test card that can detect this biomarker based on this antibody is known to those skilled in the art.

[0053] In one embodiment, detection reagents / detection reagent compositions, test strips, test cards, and / or kits can be prepared for the following detections, including but not limited to immunoassays, including competitive immunoassays, non - competitive immunoassays, enzyme - linked immunosorbent assays (ELISA), immunohistochemical assays, chemiluminescent assays, Western blot assays, and Dot blot assays; mass spectrometry assays.

[0054] The expression level of a biomarker can be detected by targeting a binding partner of the biomarker. The reagent for detecting the expression level of the biomarker can be an antibody or its derivative. In one embodiment, the expression level of the biomarker can be detected by targeting a binding partner of the biomarker; optionally, the expression level of the biomarker is detected by an antibody targeting the biomarker; optionally, the antibody is selected from monoclonal antibodies, polyclonal antibodies, antibody fragments, humanized antibodies, camel antibodies, chimeric antibodies. In one embodiment, the antibody can be further modified to carry a detectable label, such as a label that can be detected by chemiluminescence methods.

[0055] The above-mentioned immunoassay methods are known in the art, and reference can be made to (Tijssen, Practice and Theory of Enzyme Immunoassays (Elsevier, Amsterdam, 1985); Campbell, Monoclonal Antibody Technology (Elsevier, Amsterdam, 1984); Hurrell, Monoclonal Hybridoma Antibodies: Techniques and Applications (CRC Press, Boca Raton, FL, 1982); and Zola, Monoclonal Antibodies: A Manual of Techniques, pp. 147-158 (CRC Press, Inc., 1987)).

[0056] Mass spectrometry methods are also known in the art, and reference can be made to A review on mass spectrometry-based quantitative proteomics: Targeted and data independent acquisition, Anal Chim Acta. 2017 Apr 29;964:7-23.doi:10.1016 / j.aca.2017.01.059.

[0057] In one embodiment, the expression level of the biomarker is detected by a reagent capable of specifically binding to the biomarker; optionally, the expression level of the biomarker is detected by targeting a binding partner of the biomarker; optionally, the expression level of the biomarker is detected by an antibody targeting the biomarker; optionally, the antibody is selected from monoclonal antibodies, polyclonal antibodies, antibody fragments, humanized antibodies, camel antibodies, chimeric antibodies.

[0058] Methods for preparing antibodies, such as monoclonal antibodies, are known to those skilled in the art. See Clackson et al., Nature 352, 624 - 628 (1991) and Marks et al., J. Mol. Biol. 222, 581 - 597 (1991). Antibodies targeting biomarkers in the present invention are also commercially available. For example, for the detection of HER2, commercially available ELISA kits containing antibodies against HER2 can be used.

[0059] In one embodiment, the reagent can be further modified to carry a detectable label. In one embodiment, the reagent can be further modified with a label detectable by chemiluminescence methods. In one embodiment, the reagent can be an antibody modified to carry a detectable label. In one embodiment, the reagent can be an antibody modified to carry a label detectable by chemiluminescence methods. In one embodiment, detectable labels can include, for example, colloidal gold, quantum dots, horseradish peroxidase (HRP), alkaline phosphatase (AP), glucose oxidase, radioisotopes, fluorescent reporters, etc. In one embodiment, the antibody can be further labeled with biotin and / or acridinium ester. In one embodiment, the antibody can be further labeled with colloidal gold. In one embodiment, the antibody can be further labeled with quantum dots. In one embodiment, the antibody is a biotin-labeled HER-2 monoclonal antibody and an acridinium ester-labeled HER-2 monoclonal antibody.

[0060] In one embodiment, the detection reagent / detection reagent composition, test strip, test card, and / or kit can be used for chemiluminescence detection; optionally, the detection reagent / detection reagent composition, test strip, test card, and / or kit can be used in conjunction with a fully automated chemiluminescence detection instrument or an automated / digital detection platform.

[0061] In one embodiment, the subject is a subject for whom it is difficult to diagnose major depressive disorder using current mental disorder diagnosis and classification systems, such as adolescent subjects; optionally, the current mental disorder diagnosis and classification system is: a system based on the DSM of the American Psychiatric Association (such as DSM V).

[0062] In a second aspect, the present invention further provides a kit for the specific diagnosis, differentiation, and / or differential diagnosis of adolescent males suffering from major depressive disorder, prepared based on a detection reagent for HER2 protein.

[0063] In one embodiment, the kit contains streptavidin-coated magnetic microparticles, acridinium ester-labeled mouse anti-human HER-2 monoclonal antibody, and biotin-labeled mouse anti-human HER-2 monoclonal antibody.

[0064] In one embodiment, the kit further includes a calibrator, and the calibrator is HER2 protein.

[0065] In one embodiment, the kit further includes a quality control product, and the quality control product is HER2 protein.

[0066] In one embodiment, the streptavidin-coated magnetic microparticles are formulated with Tris buffer.

[0067] In one embodiment, the acridinium ester-labeled mouse anti-human HER-2 monoclonal antibody is formulated with MES buffer.

[0068] In one embodiment, the biotin-labeled mouse anti-human HER-2 monoclonal antibody is formulated with MES buffer.

[0069] In one embodiment, the HER2 protein used as a calibrator is formulated with MES buffer.

[0070] In one embodiment, the HER2 protein used as a quality control product is formulated with MES buffer.

[0071] In a third aspect, the present invention further provides a test strip for specific diagnosis, differentiation, and / or differential diagnosis of adolescent males suffering from major depressive disorder, prepared based on a detection reagent for HER2 protein.

[0072] In one embodiment, the test strip is a test strip based on colloidal gold as a marker. In one embodiment, the test strip includes: a colloidal gold pad on which a colloidal gold-labeled mouse anti-human HER-2 monoclonal antibody is sprayed; a test line on which a mouse anti-human HER-2 monoclonal antibody is coated; and a quality control line on which a goat anti-mouse polyclonal antibody or a rabbit anti-mouse polyclonal antibody or a recombinant human HER-2 antigen is coated.

[0073] In one embodiment, the test strip is a test strip based on fluorescence chromatography. In one embodiment, the test strip includes: a marker pad containing a fluorescence-labeled mouse anti-human HER-2 monoclonal antibody; a test line on which a mouse anti-human HER-2 monoclonal antibody is coated; and a quality control line on which a goat anti-mouse polyclonal antibody or a rabbit anti-mouse polyclonal antibody or a recombinant human HER-2 antigen is coated.

[0074] In one embodiment, the test strip is a test strip based on quantum dot fluorescence. In one embodiment, the test strip comprises: a marker pad containing a quantum dot-labeled mouse anti-human HER-2 monoclonal antibody; a detection line coated with a mouse anti-human HER-2 monoclonal antibody; and a quality control line coated with a goat anti-mouse polyclonal antibody or a rabbit anti-mouse polyclonal antibody or a recombinant human HER-2 antigen.

[0075] The present invention comprehensively analyzed and compared the plasma proteomes of samples from adolescents with major depressive disorder, adolescents with bipolar disorder, adolescents with schizophrenia, and healthy adolescents by DIA mass spectrometry. The results showed that the HER2 protein in samples from adolescent male and female patients with major depressive disorder showed significant differences from healthy adolescent males at the plasma proteomic level. For the purpose of clinical diagnosis application, the present invention further verified the HER2 protein obtained by mass spectrometry in independent samples by ELISA method. Subsequently, further verification was performed on the independent samples by a chemiluminescence-based sandwich immunoassay. Finally, the present invention unexpectedly found that the expression of HER2 has gender differences in adolescent males and females. The present invention determines that HER2 can be used as a biomarker with high sensitivity and specificity for clinically specifically diagnosing adolescent males with major depressive disorder. Moreover, HER2 can also be used as a biomarker for clinically differentiating and differentially diagnosing adolescent males with major depressive disorder from bipolar disorder and schizophrenia.

[0076] Examples

[0077] The following examples further illustrate the present invention by way of example. It should be understood that the present invention is not limited to any one of the following examples.

[0078] Example 1 - Enrollment Criteria and Sample Collection

[0079] 1.1 Enrollment Criteria

[0080] This study was approved by the Ethics Committee of a affiliated hospital of a certain medical university in China. Participants were included in the study if they met the following criteria: (1) aged between 10 and 18 years old, (2) diagnosed with major depressive disorder (MDD), bipolar disorder (BD), or schizophrenia (SZ) according to DSM-V, or a healthy individual matched with the disease group, (3) first onset, and MDD participants had not received drug treatment, (4) both the participants and their legal guardians provided informed consent.

[0081] The inclusion of the healthy control group (HC) was conducted by two experienced and trained psychiatric professionals based on the Structured Clinical Interview for DSM-5 (SCID-5). Meanwhile, the healthy control group was evaluated using the HAMD-17 and HAMA-14 scales.

[0082] Finally, a total of 479 subjects were recruited in this study, including 146 MDD patients (MDD group), 112 BD patients (BD group), 61 SZ patients (SC group), and 160 HC subjects (HC group).

[0083] 1.2 Sample collection

[0084] The above-mentioned included adolescents underwent blood sample collection between 8 am and 12 pm. Fasting blood samples were collected by venipuncture using EDTA-coated tubes to prevent blood clotting. The samples were sent to the laboratory for processing within 30 minutes. Subsequently, the tubes were centrifuged at 3000 rpm for 10 minutes at 4°C. After centrifugation, the plasma was carefully separated and immediately stored at -80°C until further proteomic analysis. The time from plasma collection to storage was no more than 2 hours.

[0085] Example 2 - Mass spectrometry detection and data analysis

[0086] 2.1 Sample processing for DIA analysis

[0087] High-abundance proteins were removed from plasma samples using liquid chromatography combined with a multiple affinity removal column (Agilent Technologies Inc.). Protein concentration was determined using a BCA protein assay kit (Thermo Scientific). Subsequently, proteins in the plasma were precipitated using trichloroacetic acid solution, reduced with 20 mM tris(2-carboxyethyl)phosphine hydrochloride (TCEP), and alkylated with 40 mM iodoacetamide in the dark for 30 minutes. The mixture was digested overnight with trypsin at an enzyme-to-sample protein ratio of 1:100 (w / w) at 37°C. The lysate was desalted using a Monospin C18 column (GL Science, Tokyo, Japan), and then vacuum centrifuged to dryness. The extracted peptides were redissolved in 0.1% formic acid before data-independent acquisition (DIA) analysis.

[0088] 2.2 Preparation of library samples

[0089] The extracted peptide segments were prepared as described above. The peptide segments of each sample were mixed to obtain a 100 μg purified mixture, and then dried by vacuum centrifugation. The dried peptide segments were redissolved in Milli-Q water containing 2% acetonitrile for fractionation.

[0090] 2.3 High-pH reversed-phase fractionation

[0091] The peptide fragments were analyzed on the chromatographic column (BEH C18, 1.7μm, 1mm×150mm) was fractionated on a Waters XevoTMACQUITY UPLC (Waters) column connected to a Waters XevoTMACQUITY UPLC (Waters) at a pH of 10. The eluted peptides were collected every 60 seconds, for a total of 62 fractions over a time span of 80 minutes, which were cross-merged into 31 fractions and dried by vacuum centrifugation. The dried peptides were dissolved in 10μl Mill-Q water containing 0.1% formic acid, and then iRT peptides were added for chromatographic calibration.

[0092] 2.4 Liquid chromatography

[0093] Nanoflow reversed-phase chromatography was performed using a nanoElute liquid chromatography system (Bruker Daltonics). Peptides were separated at a flow rate of 300 nl / min using a nonlinear gradient starting with 2% acetonitrile containing 0.1% formic acid, gradually increasing to 22% acetonitrile containing 0.1% formic acid, then increasing to 37% in 8 min, then further increasing to 100% in 5 min, and finally maintaining 100% acetonitrile for 7 min before re-equilibration. A homemade column (25 cm × 75 μm, 1.5 μm C18-AQ particles) was used. Mobile phases A and B were water and acetonitrile containing 0.1% formic acid, respectively.

[0094] 2.5 Mass spectrometry analysis

[0095] The LC was connected online to a timsTOF Pro (Bruker) via a CaptiveSpray nano-electrospray ion source. All 31 fraction samples were subjected to mass spectrometry analysis in data-dependent mode. The accumulation and ramp times were set to 100 ms, and mass spectra were recorded in the range of m / z 100–1700 in positive electrospray mode to generate a spectral library. Another analysis was performed in data-independent mode, including MS1 scans from m / z 400 to m / z 1200 and 64 MS2 windows, using a dia-PASEF acquisition scheme to cover the mass range of m / z 400 to m / z 1200. Ion mobility was scanned from 0.6 to 1.6 Vs / cm2. The collision energy was increased linearly as a function of mobility, from 59 eV at 1 / K0 = 1.6 Vs / cm2 to 20 eV at 1 / K0 = 0.6 Vs / cm2.

[0096] 2.6 Library generation and DIA data analysis

[0097] A spectral library was generated using Spectronaut version 15.6 (Biognosys) against the UNIPROT human database (containing only reviewed entries). All parameters were set to default. The DIA files were processed in the default mode, except that the correction factor for the XIC IM extraction window was set to 0.8.

[0098] 2.7 Parallel Reaction Monitoring Validation

[0099] For the PRM validation of the selected biomarkers, peptide precursors were scheduled within a 30-minute retention time interval using timsTOF Pro2 (Bruker). PRM quantification was performed using Skyline software.

[0100] 2.8 Machine Learning and Statistical Analysis

[0101] This study employed multiple machine learning methods (such as PLS-DA, SVM, MLP, LR, and RF) to screen for protein biomarkers to distinguish patients with MDD, BD, SZ, and HC. Missing values were imputed by the DBB strategy, and the performance of the models was evaluated using the ROC curve, AUROC value, and balanced accuracy. The study also conducted LOOCV and permutation tests, evaluated the model robustness using Q2 and R2, and analyzed the importance of proteins through VIP values. In the statistical analysis, lowess regression was used to impute and normalize the data, and gene ontology and pathway enrichment analyses were performed in combination with the GO and KEGG databases. Differentially expressed proteins (DEPs) were screened by FDR-adjusted P-values and fold changes, and protein expression clustering, interaction networks, and pathway analyses were performed using Mfuzz, Fisher's exact test, and String. Finally, the activation status of biological processes was predicted by Ingenuity Pathway Analysis and the z-score algorithm, and data visualization was performed using ggplot2 and Cytoscape.

[0102] 2.9 Determination of DEPs at the Mass Spectrometry Level

[0103] As described above, the samples were subjected to data-independent acquisition (DIA) mass spectrometry analysis, and were analyzed using trapped ion mobility spectrometry combined with time-of-flight mass spectrometry (TIMS-TOF MS) and parallel accumulation - sequential fragmentation (PASEF) methods.

[0104] After removing extreme values, the results showed that the mass spectrometry signal intensity of HER2 showed significant differences between the MDD groups and the HC groups in adolescent males and adolescent females ( Figure 1 ), while there were no significant differences between the SZ groups and the HC groups in adolescent males and adolescent females ( Figure 2) When only analyzing the mass spectrometry data of adolescent males, it was found that the mass spectrometry signal intensity of HER2 showed a significant difference between the MDD group and the HC group of adolescent males, while there was no significant difference between the BD group and the SZ group of adolescent males and the HC group. Figure 3 ) The above results suggest that the HER2 protein may be a biomarker for detecting adolescent MDD.

[0105] Example 3 - ELISA identification of differential expression of HER2 protein in MDD patients

[0106] Using the blood samples from Example 1, serum was isolated therefrom. The serum samples were from: 36 healthy adolescent males, 8 healthy adolescent females, 14 adolescent males with MDD, and 22 adolescent females with MDD. A kit (R&D Systems, #DHER20) was used to measure and compare the content of HER2 protein in the serum samples of MDD patients and healthy people from the above cohorts.

[0107] Before use, all reagents and samples were placed at room temperature. In a 96 - well antibody - coated plate, 50 μL of diluent was added, and 50 μL of standard, control, and samples were added to each corresponding well. After covering with a sealing film, it was incubated at room temperature on a horizontal microplate shaker for 2 h. The liquid in each well was aspirated and washed, and this process was repeated 3 times, for a total of 4 washes. 200 μL of human HER2 antibody and horseradish peroxidase (HRP) conjugate (R&D·Systems) were added to each well. Covered with a new sealing film. Incubated on a shaker at room temperature for 2 hours. The liquid in each well was aspirated and washed, for a total of 4 washes. Then 200 μL of substrate solution was added to each well. Incubated at room temperature for 30 minutes, protected from light. Subsequently, 50 μL of stop solution was added to terminate the reaction, and the OD450 was read within 30 min. Finally, the expression of HER2 was calculated from the OD450 readings. The measured result data are shown in Figure 4 and Figure 5 in.

[0108] The results showed that the content of HER2 in the blood of the HC group was significantly higher than that in the blood of the MDD group (6.818 ± 1.803 ng / mL vs 5.108 ± 1.870 ng / mL).

[0109] Furthermore, the samples of adolescent males and females were grouped. There was no statistically significant difference in the content of HER2 in the blood of adolescent females (6.164 ± 0.598 ng / mL vs 5.306 ± 2.000 ng / mL). In contrast, the content of HER2 in the blood of adolescent males was significantly higher in the HC group than in the MDD group (6.963 ± 1.950 ng / mL vs 4.796 ± 1.669 ng / mL). This result indicates that the expression of HER2 has gender differences in adolescent males and females. The expression of HER2 protein in adolescent male MDD patients is statistically significantly different from that in adolescent male HC, adolescent male SZ, and BD patients.

[0110] Example 4 - Verification of the Detection Reliability of HER2 as a Specific Diagnostic Marker in Adolescent Males with Major Depressive Disorder

[0111] To verify the detection reliability of HER2 as a specific diagnostic marker in adolescent males with major depressive disorder, Nanjing Renmai Biotechnology Co., Ltd. was commissioned to perform a verification experiment on the sera of 58 clinically diagnosed adolescent males with MDD and 41 healthy individuals undergoing physical examinations using a sandwich ELISA based on chemiluminescence method targeting this biomarker.

[0112] 4.1 Inclusion Criteria and Sample Collection

[0113] This study used 58 clinically diagnosed adolescent males with MDD and 41 healthy adolescent males. The inclusion criteria and sample collection of the samples were the same as those in Example 1.

[0114] 4.2 Sandwich ELISA Detection

[0115] 4.2.1 Preparation of R1 Reagent (Streptavidin-Coated Magnetic Particles)

[0116] 200 mg of carboxyl magnetic particles were resuspended in 100 mL of 0.01 mol / L 2-(N-morpholino)ethanesulfonic acid solution (MES solution), 20 mg of streptavidin (SA) was added, and the reaction was carried out at 37 °C for 30 min. Then, 2 mL of 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) at 10 mg / mL was added, and after reacting for 2 h, the mixture was adsorbed using a magnetic stand, allowed to stand for 5 min, the liquid was removed, 200 mL of 0.01 mol / L Tris buffer was added, and the above process was repeated for a total of 3 washes. Finally, it was made up to 320 mL with 0.01 mol / L Tris buffer (containing 1% w / v BSA) and stored at 2 - 8 °C.

[0117] 4.2.2 Preparation of R3 Reagent (Acridinium Ester-Labeled HER-2 Monoclonal Antibody)

[0118] The R3 reagent (acridinium ester-labeled anti-HER-2 monoclonal antibody) was purchased from Wancheng Biotechnology (Wancheng Biotechnology, product number: CW0342). The two were mixed according to a molar ratio of acridinium ester: anti-HER-2 antibody = 10:1. After reacting at 37 °C for 1 h, dialysis purification was carried out with phosphate buffer to obtain the acridinium ester-labeled anti-HER-2 antibody. It was diluted to 0.2 μg / mL with 100 mM MES buffer at pH 6.0 containing 1% w / v BSA, 0.15% v / v Tween 20, and 0.1% v / v ProClin 300 preservative.

[0119] 4.2.3 Preparation of the R4 reagent (biotin-labeled anti-HER-2 monoclonal antibody)

[0120] The R4 reagent (biotin-labeled anti-HER-2 monoclonal antibody) was purchased from Wancheng Biotechnology (Wancheng Biotechnology, product number: CW0341).

[0121] The two were mixed according to a molar ratio of biotin: anti-HER-2 antibody = 10:1. After reacting at 37 °C for 30 min, dialysis purification was carried out with phosphate buffer to obtain the biotin-labeled anti-HER-2 antibody. It was diluted to 1.0 μg / mL with 100 mM MES buffer at pH 6.0 containing 1% w / v BSA, 0.15% v / v Tween 20, 0.1% v / v ProClin 300 preservative, and 50 μg / mL blocker.

[0122] 4.2.4 Detection results of the double-antibody sandwich method based on the chemiluminescence method

[0123] Using the detection reagent prepared above based on the chemiluminescence method, it was made into the form of a corresponding kit. The kit further included calibrators and quality control products, both of which were HER-2 antigens prepared with MES buffer. 41 healthy adolescent male samples and 58 adolescent male samples with MDD were tested respectively, and a T-test was performed on the measured luminescence values. The chemiluminescence measurement was completed using the fully automatic chemiluminescence analyzer A6I (Nanjing Renmai Biotechnology Co., Ltd.). The detection results are shown in Table 1 below.

[0124] Table 1 Test results of major depressive disorder samples and healthy person samples

[0125]

[0126]

[0127] The average measured luminescence value of the major depressive disorder samples (10930) was 57.92% lower than that of the healthy person samples (25976). Further, based on the luminescence values detected above, a T-test was performed, and the P value was less than 0.0001 ( Figure 6)。Perform an ROC curve on the test results, with an AUC value of 0.919( Figure 7 )。The specificity of this result is 93% and the sensitivity is 80% (Cutoff value = 12250). The results of the chemiluminescence test once again prove that patients with major depressive disorder can be effectively identified by detecting the HER2 protein.

[0128] The present invention further includes the following technical solutions and combinations selected from any one of them:

[0129] Technical solution group A:

[0130] A1. Use of a reagent for detecting a biomarker in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for specifically diagnosing an adolescent male with major depressive disorder in the adolescent male population, wherein the kit, test strip or test card includes a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder, the biomarker includes HER2, wherein a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder, and the kit, test strip or test card further includes instructions or inserts indicating that a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder; optionally, the age of the adolescent male population is 12 - 18 years old.

[0131] A2. Use of a reagent for detecting a biomarker in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for differentiating or differentially diagnosing an adolescent male with major depressive disorder and a healthy adolescent male in the adolescent male population, wherein the kit, test strip or test card includes a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder, the biomarker includes HER2, wherein a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder, and the kit, test strip or test card further includes instructions or inserts indicating that a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder; optionally, the age of the adolescent male population is 12 - 18 years old.

[0132] Use of a reagent for detecting a biomarker in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for differentiating or differentially diagnosing adolescent males with major depressive disorder and adolescent males with schizophrenia in an adolescent male population, wherein the kit, test strip or test card comprises a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder or schizophrenia, the biomarker comprises HER2, a decrease in the expression level of the biomarker as compared to a reference value indicates that the adolescent male subject has major depressive disorder, and the kit, test strip or test card further comprises instructions for use or an insert indicating that a decrease in the expression level of the biomarker as compared to the reference value indicates that the adolescent male subject has major depressive disorder; optionally, the age of the adolescent male population is 12 - 18 years old.

[0133] Use of a reagent for detecting a biomarker in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for differentiating or differentially diagnosing adolescent males with major depressive disorder and adolescent males with bipolar disorder in an adolescent male population, wherein the kit, test strip or test card comprises a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder or bipolar disorder, the biomarker comprises HER2, a decrease in the expression level of the biomarker as compared to a reference value indicates that the adolescent male subject has major depressive disorder, and the kit, test strip or test card further comprises instructions for use or an insert indicating that a decrease in the expression level of the biomarker as compared to the reference value indicates that the adolescent male subject has major depressive disorder; optionally, the age of the adolescent male population is 12 - 18 years old.

[0134] Use of a reagent for detecting a biomarker in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for differentiating or differentially diagnosing adolescent males with major depressive disorder, adolescent males with bipolar disorder, and adolescent males with schizophrenia in an adolescent male population, wherein the kit, test strip or test card comprises a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder, the biomarker comprises HER2, wherein a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder, and the kit, test strip or test card further comprises instructions or inserts indicating that a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder; optionally, the age of the adolescent male population is 12-18 years old.

[0135] Use of a reagent for detecting a biomarker in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for excluding the possibility of an adolescent male having major depressive disorder from an adolescent male population, wherein the kit, test strip or test card comprises a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder, the biomarker comprises HER2, wherein a non-decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject does not have major depressive disorder, and the kit, test strip or test card further comprises instructions or inserts indicating that a decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject has major depressive disorder; optionally, the age of the adolescent male population is 12-18 years old.

[0136] Use of a reagent for detecting a biomarker in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for excluding the possibility of an adolescent male having major depressive disorder from adolescent males suspected of having major depressive disorder or bipolar disorder, wherein the kit, test strip or test card comprises a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder or bipolar disorder, the biomarker comprises HER2, wherein a non-decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject does not have major depressive disorder, and the kit, test strip or test card further comprises instructions or inserts indicating that a decrease in the expression level of the biomarker compared to the reference value indicates that the adolescent male subject has major depressive disorder; optionally, the age of the adolescent male population is 12-18 years old.

[0137] Use of a reagent for detecting a biomarker in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for excluding the possibility of an adolescent male having major depressive disorder from adolescent males suspected of having major depressive disorder or schizophrenia, wherein the kit, test strip or test card comprises a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder or schizophrenia, the biomarker comprises HER2, wherein a non-decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject does not have major depressive disorder, and the kit, test strip or test card further comprises instructions or inserts indicating that a decrease in the expression level of the biomarker compared to the reference value indicates that the adolescent male subject has major depressive disorder; optionally, the age of the adolescent male population is 12-18 years old.

[0138] Use of a reagent for detecting a biomarker in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for excluding the possibility of an adolescent male having major depressive disorder from adolescent males suspected of having major depressive disorder, bipolar disorder or schizophrenia, wherein the kit, test strip or test card comprises a reagent for detecting the expression level of a biomarker in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder, the biomarker comprises HER2, and wherein a non-decrease in the expression level of the biomarker compared to a reference value indicates that the adolescent male subject does not have major depressive disorder, and the kit, test strip or test card further comprises instructions for use or an insert indicating that a decrease in the expression level of the biomarker compared to the reference value indicates that the adolescent male subject has major depressive disorder; optionally, the age of the adolescent male population is 12-18 years old.

[0139] A10. The use according to any one of technical solutions A1-A9, wherein the kit, test strip or test card further comprises: a calibrator and / or a calibrator, and the calibrator and / or the calibrator is HER-2 protein.

[0140] A11. The use according to any one of technical solutions A1-A10, wherein the sample is a blood sample, preferably the sample is a plasma or serum sample.

[0141] A12. The use according to any one of technical solutions A1-A11, wherein the reference value is the expression level of the biomarker in a sample from a healthy subject or from a standard sample; optionally, the reference value is the concentration of HER2 protein in a sample from a healthy subject or from a standard sample.

[0142] A13. The use according to any one of technical solutions A1-A12, wherein the expression level of the biomarker is determined by a method selected from the following: immunoassay, including competitive immunoassay, non-competitive immunoassay, enzyme-linked immunosorbent assay (ELISA), immunohistochemical assay, chemiluminescence assay, Western blot assay and Eot blot assay; mass spectrometry assay.

[0143] A14. The use according to any one of technical solutions A1-A13, wherein the expression level of the biomarker is detected by a reagent capable of specifically binding to the biomarker; optionally, the expression level of the biomarker is detected by a binding partner targeting the biomarker; optionally, the expression level of the biomarker is detected by an antibody targeting the biomarker; optionally, the antibody is selected from monoclonal antibody, polyclonal antibody, antibody fragment, humanized antibody, camel antibody, chimeric antibody.

[0144] A15. The use according to any one of technical solutions A1 - A14, wherein the reagent can be further modified to carry a detectable label; optionally, the antibody can be further modified to carry a detectable label, for example, a label detectable by chemiluminescence method; optionally, the antibody can be further biotinylated and / or acridinium ester labeled; optionally, the antibody is a biotinylated anti-HER-2 monoclonal antibody and an acridinium ester labeled anti-HER-2 monoclonal antibody.

[0145] A16. The use according to any one of technical solutions A1 - A15, wherein the subject is a subject in whom it is difficult to diagnose major depressive disorder by the current mental disorder diagnosis and classification system; optionally, the current mental disorder diagnosis and classification system is: a system based on the DSM of the American Psychiatric Association (such as DSM V).

[0146] A17. The use according to any one of technical solutions A1 - A16, wherein the kit, test strip or test card can be used for chemiluminescence detection.

[0147] A18. The use according to any one of technical solutions A1 - A17, wherein the kit, test strip or test card can be used in combination with a fully automatic chemiluminescence detection instrument or an automated / digital detection platform.

[0148] A19. The use according to any one of technical solutions A1 - A18, wherein the kit, test strip or test card comprises: streptavidin-coated magnetic particles, biotinylated anti-HER-2 monoclonal antibody, and acridinium ester labeled anti-HER-2 monoclonal antibody.

[0149] A20. The use according to technical solution A19, wherein the kit, test strip or test card further comprises: a calibrator and / or a calibrator, and the calibrator and / or the calibrator is HER-2 protein.

[0150] Technical solution group B:

[0151] B1. A kit for specific diagnosis, differentiation and / or differential diagnosis of adolescent males with major depressive disorder, comprising the following components: streptavidin-coated magnetic particles, acridinium ester labeled mouse anti-human HER-2 monoclonal antibody, and biotinylated mouse anti-human HER-2 monoclonal antibody.

[0152] B2. A kit for detecting the concentration of HER2 protein, comprising the following components: streptavidin-coated magnetic particles, acridinium ester labeled mouse anti-human HER-2 monoclonal antibody, and biotinylated mouse anti-human HER-2 monoclonal antibody.

[0153] Kit according to technical solution B1 or B2, wherein the kit further comprises a calibrator which is HER2 protein; and / or the kit further comprises a quality control product which is HER2 protein.

[0154] B4. The kit according to any one of technical solutions B1 - B3, wherein

[0155] the streptavidin - coated magnetic microparticles are prepared with Tris buffer; and / or

[0156] the acridinium - ester - labeled mouse anti - human HER - 2 monoclonal antibody is prepared with MES buffer; and / or

[0157] the biotin - labeled mouse anti - human HER - 2 monoclonal antibody is prepared with MES buffer; and / or

[0158] the HER2 protein used as a calibrator is prepared with MES buffer; and / or

[0159] the HER2 protein used as a quality control product is prepared with MES buffer.

[0160] B5. The kit according to any one of technical solutions B1 - B4, wherein the kit further comprises an instruction manual for indicating that when the HER - 2 concentration in a sample from an adolescent male subject detected is lower than the reference value, it is indicated that the adolescent male subject has major depressive disorder.

[0161] Technical solution group C:

[0162] C1. A test strip for specific diagnosis, differentiation and / or differential diagnosis of adolescent males with major depressive disorder, comprising the following structure:

[0163] A colloidal gold pad on which a colloidal gold - labeled mouse anti - human HER - 2 monoclonal antibody is sprayed;

[0164] A test line on which a mouse anti - human HER - 2 monoclonal antibody is coated; and

[0165] A quality control line on which a goat anti - mouse polyclonal antibody or a rabbit anti - mouse polyclonal antibody or a recombinant human HER - 2 antigen is coated.

[0166] C2. A test strip for specific diagnosis, differentiation and / or differential diagnosis of adolescent males with major depressive disorder, comprising the following structure:

[0167] A marker pad which contains a fluorescence - labeled mouse anti - human HER - 2 monoclonal antibody;

[0168] The test line, on which a mouse anti-human HER-2 monoclonal antibody is coated; and

[0169] The quality control line, on which a goat anti-mouse polyclonal antibody or a rabbit anti-mouse polyclonal antibody or a recombinant human HER-2 antigen is coated.

[0170] C3. A test strip for specific diagnosis, differentiation and / or differential diagnosis of adolescent males with major depressive disorder, comprising the following structure:

[0171] The marker pad, which contains a quantum dot-labeled mouse anti-human HER-2 monoclonal antibody;

[0172] The test line, on which a mouse anti-human HER-2 monoclonal antibody is coated; and

[0173] The quality control line, on which a goat anti-mouse polyclonal antibody or a rabbit anti-mouse polyclonal antibody or a recombinant human HER-2 antigen is coated.

[0174] C4. A test strip for detecting the concentration of HER2 protein, comprising the following structure:

[0175] The colloidal gold pad, on which a colloidal gold-labeled mouse anti-human HER-2 monoclonal antibody is sprayed;

[0176] The test line, on which a mouse anti-human HER-2 monoclonal antibody is coated; and

[0177] The quality control line, on which a goat anti-mouse polyclonal antibody or a rabbit anti-mouse polyclonal antibody or a recombinant human HER-2 antigen is coated.

[0178] C5. A test strip for detecting the concentration of HER2 protein, comprising the following structure:

[0179] The marker pad, which contains a fluorescence-labeled mouse anti-human HER-2 monoclonal antibody;

[0180] The test line, on which a mouse anti-human HER-2 monoclonal antibody is coated; and

[0181] The quality control line, on which a goat anti-mouse polyclonal antibody or a rabbit anti-mouse polyclonal antibody or a recombinant human HER-2 antigen is coated.

[0182] C6. A test strip for detecting the concentration of HER2 protein, comprising the following structure:

[0183] The marker pad, which contains a quantum dot-labeled mouse anti-human HER-2 monoclonal antibody;

[0184] The test line, on which a mouse anti-human HER-2 monoclonal antibody is coated; and

[0185] Quality control line, on which polyclonal goat anti-mouse antibody or polyclonal rabbit anti-mouse antibody or recombinant human HER-2 antigen is coated.

[0186] C7. The test strip according to any one of technical solutions C1-C6, wherein the test strip further includes an instruction manual for indicating that when the HER-2 concentration in the sample from an adolescent male subject detected is lower than the reference value, it is indicated that the adolescent male subject has major depressive disorder.

Claims

1. Use of a reagent for detecting the concentration of HER2 in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for specifically diagnosing adolescent males with major depressive disorder in the adolescent male population, wherein the kit, test strip or test card comprises a reagent for detecting the concentration of HER2 in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder, and wherein the age of the adolescent male population is 12 - 18 years old.

2. Use of a reagent for detecting the concentration of HER2 in a sample from an adolescent male subject in the preparation of a kit, test strip or test card for differentiating or differentially diagnosing adolescent males with major depressive disorder, adolescent males with bipolar disorder, and adolescent males with schizophrenia in the adolescent male population, wherein the kit, test strip or test card comprises a reagent for detecting the concentration of HER2 in a sample from an adolescent male subject, wherein the adolescent male subject is suspected of having major depressive disorder, and wherein the age of the adolescent male population is 12 - 18 years old.

3. Use according to any one of claims 1 - 2, wherein the sample is blood, plasma or serum.

4. Use according to any one of claims 1 - 3, wherein the HER2 concentration is determined by chemiluminescence.

5. Use according to any one of claims 1 - 4, wherein the reagent for detecting the concentration of HER2 in a sample from an adolescent male subject is detected by an antibody capable of specifically binding to HER2; optionally, the antibody is selected from monoclonal antibodies or polyclonal antibodies.

6. Use according to any one of claims 1 - 5, wherein the antibody is a biotin-labeled HER-2 monoclonal antibody and an acridinium ester-labeled HER-2 monoclonal antibody.

7. Use according to any one of claims 1-6, wherein the kit, test strip or test card further comprises: Calibrator and / or calibrator, the calibrator and / or calibrator being HER-2 protein.

8. Use according to any one of claims 1 - 7, wherein the subject is a subject in whom it is difficult to diagnose major depressive disorder by the system of DSM V of the American Psychiatric Association.

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