24 results about "Zona pellucida" patented technology

Provided herein are methods for coating a microparticle, cell, or therapeutic agent with a biomimetic zona pellucida (BZP). Also provided herein are BZP-coated cells prepared using the enclosed methods. The methods for using the BZP-coated microparticle to treat diabetes are also provided.

An automated method for assessing the ZP binding capacity of sperm from morphological characteristics of sperm in assisted reproduction using deep learning is disclosed. The invention also provides a method for predicting fertilization success based on ZP binding capacity of sperm in clinical assisted reproduction.

The invention discloses a dairy cow early embryo sex identification method, and belongs to the technical field of dairy cow breeding and breeding. The identification method comprises the following steps: (1) culturing a dairy cow embryo to an early blastocyst stage, starting to perform single-embryo single-droplet culture, and after culturing until a blastocyst cavity is formed, punching a blastocyst zona pellucida to promote the blastocyst to shrink and release blastocyst cavity liquid; (2) collecting blastocyst cavity liquid, and extracting embryo free DNA in the blastocyst cavity liquid; (3) by taking the free DNA as a template, designing a primer to amplify a specific repetitive sequence ChrY fragment of the bull Y chromosome; if the amplification product exists, the embryo is identified as a bull, and if the amplification product does not exist, the embryo is identified as a cow. The method can ensure that the embryo free DNA in the obtained blastocyst cavity fluid is only derived from the embryo, and can effectively avoid embryo injury. According to the method disclosed by the invention, the copy number of the amplified target area in the bull Y chromosome is more than 2000 times, a detection signal can be amplified, and the detection accuracy is improved.

This invention relates to the field of bioengineering technology, and in particular to a method for simultaneously collecting the inner cell mass and trophectoderm of mammalian blastocysts. The method includes the following steps: (1) after removing the zona pellucida of the blastocyst, the blastocyst is cut into an inner cell mass and a trophectoderm portion, and the cut trophectoderm portion is transferred to a culture medium for collection; (2) the inner cell mass portion obtained in step (1) is sequentially placed in an antibody treatment solution and a complement treatment solution for incubation; (3) the inner cell mass portion treated in step (2) is pipetted to obtain a purified inner cell mass. This method achieves simultaneous and efficient separation of two key cell lines from the blastocyst, fundamentally changing the traditional method's "resource waste" model of only obtaining the inner cell mass while discarding the trophectoderm, and providing more comprehensive experimental materials for embryonic development, placental biology, and stem cell research.

This invention relates to the field of reproductive engineering technology, specifically to the application of rutabagastone in improving the cryopreservation and in vitro maturation of oocytes. This invention is the first to clearly demonstrate that rutabagastone can improve the quality of cryopreservation and in vitro maturation of frozen oocytes. By adding rutabagastone to the cryopreservation solution, thawing solution, and in vitro maturation solution, this invention reduces cryopreservation damage to oocytes, resulting in increased survival rate after thawing, improved mitochondrial function, reduced damage to the oocyte spindle and zona pellucida, reduced oxidative stress levels, and ultimately improved subsequent developmental capacity of frozen oocytes.

The utility model provides a laser-assisted zona pellucida punching system, and belongs to the technical field of medical instruments. The system comprises a laser emission module, a guiding device and a microscope assembly, the laser emission module is arranged beside the microscope assembly and comprises a laser light source and an optical fiber collimator which are connected in sequence, the output end of the laser emission module faces the guiding device, and the laser emission module is configured to be capable of two-dimensionally moving relative to the light incident side of the guiding device. The guiding device comprises a concave reflector, and the concave reflector is arranged at an angle relative to the light emitting direction of the optical fiber collimator. The microscope assembly comprises a secondary reflector, an objective lens and a culture dish which are arranged in sequence, and the secondary reflector is arranged at the incident end of the microscope assembly and used for reflecting the incident light beam through the guide device to the objective lens and focusing the light beam on the culture dish. By the adoption of the system and the punching method, the adverse effect caused by the fact that an arc guide rail is adopted for conducting laser transparent belt punching operation in the related technology can be avoided, and the punching precision is improved.

The invention relates to the field of cell genetic engineering, and discloses a high-sensitivity embryo chromatin analysis method which comprises the following steps: S1, puncturing an embryo zona pellucida through a micromanipulation instrument, sucking 5-8 trophoblast cells, and immersing a sample into 4 DEG C chromatin stabilizing liquid containing 1mM of ethylenediamine tetraacetic acid and bovine serum albumin within 8 seconds after the sample is obtained; s2, processing the sample by adopting a gradient fixation method; s3, sequentially carrying out low-intensity ultrasonication and restriction enzyme combined treatment; s4, carrying out specific labeling by using a double-labeling probe system; s5, implementing two-stage signal amplification in the micro-fluidic chip; and S6, carrying out three-dimensional space conformation analysis by adopting a super-resolution microscope. A silicon nitride microscopic needle penetrates through the zona pellucida at a puncture angle of 75-85 degrees, and the suction speed is controlled to be 0.5 mu L / s in combination with a PID closed loop, so that the DNA fragmentation rate can be reduced to 2.1 + / -0.3% (the proportion of 5kb fragments is 98.5%), and the embryo survival rate is gt; the H3K9 acetylation modification level is improved by 5.3 times, and the chromatin accessibility is improved.

The invention discloses a carnitine micromolecule mediated metabolism regulation-based sheep non-reproductive season oestrus accurate induction method, and belongs to the technical field of livestock breeding. According to the method, through supplementary feeding of high-purity L-carnitine (larger than or equal to 99%, and the dosage is 200 mg / kg BW), a three-level regulation and control network of metabolism reprogramming (AMPK / mTOR pathway activation)-epigenetic remodeling (CYP19A1 promoter demethylation)-neuroendocrine activation (Kisspeptin / GnRH axis) is constructed, and the estrus rate of the ewes in the non-breeding season is larger than or equal to 50% (the control group is 3%, Plt is 0.01). The method specifically comprises the following steps: continuously intervening for 90 days by oral administration of a DL-carnitine solution, detecting dynamic changes of serum E2 (peak value is greater than or equal to 868pg / mL) and LH (peak value is greater than or equal to 45ng / mL) by ELISA (enzyme-linked immuno sorbent assay), and confirming ovarian follicular development (clear zona pellucida structure) by HE dyeing. The traditional hormone dependence mode is innovatively broken through, the inflammation risk of the reproductive system is avoided, and a new non-hormone intervention strategy is provided for the ruminant seasonal reproductive disorder.

The invention relates to a mitochondrial delivery system and a preparation method and application thereof. According to the mitochondrial delivery system, safe and efficient cell entry of active mitochondria can be achieved, the mitochondrial delivery system has appropriate particle size and surface potential, toxicity to cells is avoided while the entrapment rate of the mitochondria is effectively improved and the electrochemical stability and biological activity of a mitochondrial membrane are guaranteed, and the mitochondrial cell entry efficiency is remarkably improved. The mitochondrial delivery system not only can transplant active mitochondria into common cells to repair cell damage, but also can transplant the active mitochondria into egg cells to play a role across a zona pellucida and a plasma membrane barrier. The mitochondrial delivery system can be used for treating mitochondrial diseases related to mitochondrial dysfunction, especially infertility caused by mitochondrial dysfunction of egg cells.

The invention discloses a mammal in-vitro semen optimization method based on charge treatment, and relates to the technical field of livestock breeding biology, and the method comprises the following steps: (a) obtaining mammal semen in an in-vitro environment; (b) putting the sperm into a charge treatment device, and applying a directional electric field to enable the surface of the sperm to carry charges with preset polarity; (c) controlling the electric field intensity to be 5mV and the treatment time to be 30s-60s, so that the sperms obtain the charge attribute opposite to the surface charge of the ovum zona pellucida; (d) directly applying the treated seminal fluid to artificial insemination or in-vitro insemination operation, the technology can improve the success rate that the sperms penetrate through zona pellucida, especially has a breakthrough improvement effect on pathological infertility caused by unbalanced reproductive nutrition of female animals, and is suitable for artificial insemination and embryo transplantation scenes of various mammals at the same time; and high-quality breeding stock resources are efficiently utilized on the premise that the operation convenience is guaranteed.

The application belongs to the technical field of biological medicine, and particularly relates to a drug carrier targeting ovaries and a preparation method and application thereof. Specifically, the application provides a drug carrier ZIF-8-GH@ZP3 targeting ovaries, so that targeted treatment for POI is realized, growth hormone is released in a targeted manner to improve the treatment effect and reduce the adverse reactions of growth hormone. The drug carrier is designed by using a zeolite imidazole framework (ZIF-8) and a zona pellucida antibody (ZP3Ab), can release growth hormone in a targeted manner according to different levels of developing follicles in the ovaries, based on the dual functions of targeting and promoting the development of dominant follicles, finally realizes targeted treatment for primary ovarian insufficiency, and has good practical application value.

The application discloses a kind of high efficiency gentle mammal blastocyst single cell dissociation method.The method includes the following steps: (1) zona pellucida removal: the blastocyst is placed in calcium magnesium table type liquid, and zona pellucida is completely removed by dissolving and physical blowing operation;(2) enzyme digestion incubation: the blastocyst after step (1) is handled and is transferred to 35-37 ℃ pre-equilibrated complex dissociation solution and incubated for 15-30 minutes, during which intermittent blowing is carried out, until the blastocyst is completely dissociated into single cell;(3) washing: the single cell obtained by dissociation is washed using dPBS, and the residual dissociation solution is removed.The method of the application has high efficiency, high activity and simplified operation.The dissociation process is simplified from 3-4 steps to two steps (zona pellucida dissolution+enzyme digestion incubation), the total time is shortened from more than 90 minutes to less than 30 minutes, the dissociation efficiency is significantly improved;Cell viability is increased from about 70% to more than 90%, the cell junction is gently lysed, and cell damage is reduced.

The present application belongs to the technical field of pharmaceutical preparations, and specifically relates to the new use of Tongren Wuji Baifeng Pills and a pharmaceutical composition in the preparation of a drug for treating premature ovarian failure, a pharmaceutical preparation comprising Tongren Wuji Baifeng Pills, a method for preparing same, and a method for preparing a drug for treating premature ovarian failure. In the present application, a rat model of chemotherapy-induced premature ovarian failure via intraperitoneal injection of cisplatin and a mouse model of immune-mediated premature ovarian failure via multi-point subcutaneous injection of mouse zona pellucida peptide 3 (pZP3) are established. By means of observing the effects of Tongren Wuji Baifeng Pills on the estrous cycles, organ coefficients, serum sex hormone levels, and ovarian histopathological changes in animals in both models, the therapeutic effect of Tongren Wuji Baifeng Pills on premature ovarian failure is verified and clarified. This expands the new clinical use of the drug and provides a new therapeutic drug and means for treating premature ovarian failure, thereby offering technical support for clinical precision medication.

Provided herein are methods for improving the efficiency of in vitro fertilization of animals. The method comprises (a) obtaining an oocyte from the animal, wherein the oocyte comprises an oocyte body and a zona pellucida; (b) obtaining sperm from the animal; (c) maturing the oocytes in a maturing medium; (d) removing cumulus cells from the oocytes; (e) drilling in the zona pellucida of the oocyte with a laser; (f) contacting the oocyte with the sperm; (g) co-incubating the oocytes with sperms, whereby the incubation enables in vitro fertilization of the oocytes and sperms to produce animal embryos, and whereby drilling is performed in the zona pellucida of the oocytes by laser to improve the efficiency of in vitro fertilization of the production of animal embryos.

Methods for automated ICSI include holding an oocyte in a holding device, moving an injection pipette, and positioning the oocyte to the motor position associated with the image in which the oocyte is most in focus. The methods also include using AI to: detect and determine the thickness of a zona pellucida of an oocyte, determine an image in which the oocyte is most in focus, align the oocyte and the injection pipette, label morphological structure(s) of the oocyte, and create a trajectory for the injection pipette. The methods also include perforating (by laser or piezo) the zona pellucida and injecting a spermatozoon along the injection trajectory into the oocyte.

The present invention is directed to a method for determining suitability of an oocyte post intracytoplasmic sperm injection (ICSI) for embryo implantation including determining a ratio (q) between thickness of a zona pellucida (ZP) of the oocyte and an outer diameter of the oocyte, wherein a q value between a predetermined range is indicative of an embryo developed from the oocyte having increased implantation probability. The method further includes selecting the oocyte for implantation.

A method for automated ICSI includes receiving at least one droplet containing an egg in a dish placed on a stage. The method includes using an artificial intelligence / machine learning system (AI / ML system) and an imaging system to detect a zona pellucida. The imaging system includes a microscopy system, a camera system, and a lighting system. The method includes holding the egg using a robotic microtool and lowering a robotic pipettor into the droplet. The method includes using the AI / ML system and imaging system to determine an area at which to hold the egg and positioning the robotic microtool to that area. The method includes using the AI / ML system and imaging system to instruct the robotic microtool to apply negative pressure to hold the egg to the robotic pipettor. The method includes using the AI / ML system and imaging system to determine a target location where zona ablation should be performed.

The invention discloses a B cell epitope peptide hybridoma cell strain on migratory locust zona pellucida membrane protein LmPio and a monoclonal antibody and application thereof, LH10 is inserted into a passenger-carrying structural domain of a salmonella V-type secretion system MisL and is introduced into escherichia coli DH5alpha, the LH10 is displayed and expressed on the surface of DH5alpha thallus, the hybridoma cell strain is used as an immunogen, and the hybridoma cell strain can be used for detecting the B cell epitope peptide on the migratory locust zona pellucida membrane protein LmPio. After a mouse is immunized, splenocytes of the mouse are fused with Sp2 / 0 to obtain the LH10-resistant mAb hybridoma cell. The LH10 is inserted into a salmonella peg pilus operon and is introduced into an inert carrier bacterium S9H, the LH10 is subjected to functional display expression on the surface of an S9H thallus, and an S9H-peg-LH10-antibody direct mediated agglutination test established based on the display expression on the surface of the S9H thallus can specifically recognize an anti-LH10 mAb in a hybridoma cell supernatant, so that the monoclonal antibody aiming at the B cell epitope peptide LH10 on the membrane protein LmPio can be quickly screened out.

The application relates to an image recognition-based method for positioning an oocyte polar body in an intracytoplasmic sperm injection, obtaining a microscopic digital image of an oocyte; determining an inner edge of a zona pellucida, an outer edge of a cytoplasm and a peripheral gap region located between the two according to the microscopic digital image; extracting a candidate region in the peripheral gap region; respectively judging whether each candidate region is located in the peripheral gap region, is not connected with the cytoplasm region and is not connected with the zona pellucida outer side region; determining a valid candidate region which is located in the peripheral gap region, is not connected with the cytoplasm region and is not connected with the zona pellucida outer side region; determining a target polar body region according to the valid candidate region; determining a polar body center position and polar body orientation information according to the target polar body region, and outputting a polar body positioning result. The method can be implemented based on a microscopic digital image, and is favorable for improving the stability, consistency and automation application value of the polar body positioning result.

The invention relates to the technical field of biological medicines, in particular to a construction method of Gjb2 homozygous mutation deafness mice. Injecting the Gjb2 homozygous mutant mouse embryonic stem cells into a diploid mouse embryo through zona pellucida to prepare a reconstructed embryo; and transferring the reconstructed embryo to a pseudo-pregnant female mouse, so that the postpartum generation mouse is the Gjb2 homozygous mutation deafness mouse. By adopting the construction method disclosed by the invention, the survival ratio of the mouse with homozygous mutation deafness is high, a large number of important mouse models can be provided for follow-up disease treatment exploration, and an important foundation is laid for efficient establishment of mouse models with other different mutations of GJB2.

The invention belongs to the technical field of assisted reproduction, and discloses an IVM serum-free culture solution based on fetuin. The IVM serum-free culture solution comprises the following components: an IVM serum-free culture basic solution, 0-10 mg / mL of non-typing fetuin, and the non-typing fetuin comprises fetuin-A (fetuin-A) and fetuin-B (fetuin-B), and the IVM serum-free culture basic solution comprises the IVM serum-free culture basic solution and the non-typing fetuin-A (fetuin-A) and the fetuin-B (fetuin-B), the IVM serum-free culture basic solution is prepared from 2.5 to 10 mM of sodium pyruvate, 5 mg / mL of human albumin, 10 ng / mL of epidermal growth factor (EGF), 0.075 IU / mL of follicle stimulating hormone (FSH), 0.075 IU / mL of human chorionic gonadotropin (hCG), an amino acid composition, vitamins and inorganic salt. Experiments show that by adopting the formula, the maturation rate of oocytes can reach 95.55% + / -0.51%, the natural fertilization rate can reach 90.68% + / -1.12%, the blastocyst rate can reach 85.02 + / -1.85%, the problems of cumulus expansion abnormity, zona pellucida sclerosis (ZP2 abnormal cutting), embryo morphology abnormity and the like in a serum-free system can be synchronously solved, the effect is superior to that of a single serum-B (10 [mu] g / mL) culture solution, a single serum-A (0.9 mg / mL) culture solution or a serum-containing culture solution, and serum pathogen risks and ICSI operation dependence are avoided.

The present invention provides a method for improving the efficiency of in vitro fertilization in animals. The method comprises: (a) obtaining oocytes from the animal, wherein the oocytes include the oocyte body and the zona pellucida; (b) obtaining sperm from the animal; (c) maturing the oocytes in a maturation medium; (d) removing cumulus cells from the oocytes; (e) creating holes in the zona pellucida of the oocytes with a laser; (f) contacting the oocytes with the sperm; and (g) incubating the oocytes with the sperm, wherein the incubation enables in vitro fertilization of the oocytes and sperm, and an animal embryo is produced; and the efficiency of in vitro fertilization for producing an animal embryo is improved by creating holes in the zona pellucida of the oocytes with a laser. In certain embodiments, the cumulus cells are removed from the oocytes by treatment with hyaluronidase, pipetting, and / or vortexing. In one embodiment, the oocytes are placed on a microscope stage prior to step (e).

The invention provides a dyeing method of a dwarf fertilized ovum cortex, which effectively solves the technical problems that dyes are difficult to penetrate through a fertilized ovum zona pellucida and the dyeing effect is poor in a traditional method by optimally designing a streptavidin membrane removing liquid pretreatment system and combining a fluorescence labeling spanishneedles cyclic peptide dyeing technology. After the fertilized ovum of the dwarf is obtained by inducing parturition, the efficient coloring of the cortex layer of the fertilized ovum of the dwarf can be clearly observed under a laser confocal microscope through fixing, membrane removal, permeation and dyeing treatment, and the effect is obviously superior to that of a traditional trypsin treatment method. The method is easy and convenient to operate and high in dyeing efficiency, and the leather layer dyeing effect is remarkably enhanced while the integrity of the morphological structure of the leather layer is ensured. The invention provides a reliable technical means for researching the cortex change of the bivalve fertilized egg cells in the development process, and has important application value and popularization potential.

Embodiments described herein use deep learning to automate measurement of key morphokinetic embryo features associated with viability and quality, in particular those relevant for clinical in-vitro fertilization (IVF). Systems and methods may, for example, acquire one or more digital images of one or more embryos; select one or more embryos in each digital image; and for each selected embryo, (i) computationally characterize the zona pellucida, detect the degree of fragmentation in the embryo, and for each embryo with a low fragmentation score, computationally classify the embryo's developmental stage based on whether cells constituting the embryo exceed a threshold number (e.g., nine). For an embryo consisting of a single cell, pronuclei may be detected and counted. Based on these measurements, a viability score may be assigned to the embryo.