Sperm Antibody ELISA Kit

Name: Sperm Antibody ELISA Kit
SKU: DEIA1923
Rating: 5 (1 reviews)

Regulatory status: For research use only, not for use in diagnostic procedures.

COA

SDS

Datasheet

Summary

Sample

seminal plasma

Species Reactivity

Human

Detection Method

sELISA

Intended Use

The Sperm Antibody ELISA is a reliable and quantitative test for the determination antibodies directed against human spermatozoa. This test is intended for the use with seminal plasma.
Please note: the terms "anti-spermatozoa antibodies", "anti-sperm antibodies" and "sperm antibodies" are equivalent. In these instructions the rather unwieldy but correct term "anti-spermatozoa antibodies" is used.

Contents of Kit

1. Microtiter strips coated with sperm antigen 96 wells
2. Sperm Antibody ELISA standard set - per vial 0.5 ml
Standard 1 ( 31 U/ml – colourless screw cap)
Standard 2 ( 62 U/ml – white screw cap)
Standard 3 (125 U/ml – yellow screw cap)
Standard 4 (250 U/ml – blue screw cap)
3. Control (green screw cap) equivalent to 70-120 U/ml 0.5 ml
4. Dilution Buffer (also used as blank / zero standard / 0 U/ml ) 50 ml
5. Wash Solution (10× concentrated) 50 ml
6. Enzyme Conjugate (ready for use) 8 ml
7. Substrate solution (solution of TMB, ready for use) 13 ml
8. Stop Solution (0.25 mol/l H₂SO₄) 13 ml
9. Holder for single strips 1×
10. Adhesive Sheet 2×

Storage

1. Store the reagents at 2°C-8°C (36°F-46°F).
2. The reagents remain stable until the expiration date of the kit.
3. The diluted washing solution is stable for 4 weeks at refrigerator temperatures (4°C-8°C/39°F-46°F).
4. Put caps back on the vials immediately after use.
5. Store the microtiter strips in a dry bag with desiccants. The remaining strips must be stored in the tightly resealed bag together with the desiccants. Under these storage conditions, they are stable at least for 4 weeks after opening of the sealed bag.

Precision

1. Intraassay variation coefficient: 6.44% (5.69-7.92%)
For the determination of the intraassay variation coefficient 6 kits from 6 different batches (produced on different days) were used. One patient sample (optical density about 1.0) was applied 96 times per testing procedure.
2. Interassay variation coefficient: 7.15% (6.04-8.21)
For the determination of the interassay variation coefficient one strip each of 12 kits stemming from 6 different batches (produced on different days) were used. One patient sample (optical density about 1.0) was applied 72 times per testing procedure.

General Description

Antibodies directed against spermatozoa antigens may cause infertility in women or men. The application of the Anti-Spermatozoa Antibody ELISA from CD is recommended for the diagnosis of immunologically caused disorders of fertility. Unwanted childlessness is a growing problem with which up to 20% of all couples in the reproductive age are confronted temporarily or long-term. In 20% of these cases the presence of anti-spermatozoa antibodies in the male or the female patient is detectable. The definition of infertility according to the WHO (WHO Laboratory Manual for the Examination of Human Semen and Semen Cervical-Mucus Interaction, 1999) is the absence of a conception within 12 months of unprotected intercourse. The main cause of an immunological fertility disorder is the formation of antibodies directed against spermatozoa antigens. Anti-spermatozoa antibodies exert heterogeneous effects on the ability of spermatozoa to fertilize. The inhibiting effect of anti-spermatozoa antibodies on the motility of spermatozoa by binding to their surface and by agglutinating processes is well-known. The penetration of the spermatozoa into the cervical mucus is impaired by the presence of anti-spermatozoa antibodies in the seminal plasma and/or in the cervical mucus. Anti-spermatozoa antibodies negatively influence the capacitation and the acrosome reaction of spermatozoa and thereby impede the interaction of the spermatozoa with the oocyte. The interaction of the spermatozoon with the oocyte and the subsequent binding to and penetration of the zona pellucida may be inhibited by anti-spermatozoa antibodies. The following fusion of the oocyte and a spermatozoon may also be impaired by the presence of anti-spermatozoa antibodies. The rate of pregnancies in couples with anti-spermatozoa antibodies on the part of the man or the woman are 38% lower compared to the control groups. Furthermore, an influence on the implantation and on the early embryological development could be confirmed. Anti-spermatozoa antibodies may occur dissolved in the ejaculate or bound to the surface of spermatozoa. Anti-spermatozoa antibodies may be found in men and in women. In women anti-spermatozoa antibodies may be found in cervical mucus, oviduct liquid and follicular liquid. Men having more than 50% of their spermatozoa coated with anti-spermatozoa antibodies show a conspicuously reduced rate of fertility.

Citations

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Background

Antisperm antibodies are immune antibodies that can inhibit sperm motility and weaken sperm fertilization ability. Antisperm antibodies include four types: IgG, IgA, IgM, and IgE. Antisperm antibodies in semen almost all belong to two types of immunoglobulins: IgA and IgG. They may exist in the serum and plasma of patients, in the cervical mucus of women, and on the surface of sperm in men. Whether men or women, the presence of antisperm antibodies in the body may lead to infertility. Under normal circumstances, there are no antisperm antibodies in men and women, but in some special cases, the female body's "self-defense" against sperm and semen antigens will cause the immune system to produce antibodies. Why do men have an immune response to their own sperm? The immune tolerance theory believes that when an effective antigen comes into contact with immune-active cells during the embryonic period, the immune-active cells against the antigen can be suppressed. Therefore, the individual no longer has an immune response to the antigen after adulthood. Since there is no sperm production during the embryonic period, sperm begins to be produced by the testicles during puberty, which is far beyond the period when the body establishes tolerance to itself. Therefore, the male body has not established immune tolerance to its own sperm. Therefore, sperm-specific antigens are self-antigens, or "hidden" antigens. Under normal circumstances, the antigenic substances of spermatogenic cells will not enter the blood circulation and cause autoimmune reactions. In certain pathological conditions, they can lead to anti-sperm immune reactions.

Figure 1. Production of antisperm antibodies and the mechanisms of action in the female reproductive tract that lead to infertility. (Sources: A S V, Dhama K, et al. 2019)

There are many reasons for the production of male anti-sperm antibodies, mainly including the following three: 1. Destruction of the blood-testis barrier; Under normal circumstances, the blood-testis barrier prevents sperm from contacting the body's immune system, becoming a hidden antigen and not causing an autoimmune response. When the blood-testis barrier is destroyed due to surgery, trauma, infection, etc., sperm leaks out or macrophages enter the reproductive tract to phagocytose and digest sperm cells. The sperm antigens they carry activate the immune system and produce anti-sperm antibodies. Common clinical causes include vasectomy, vas deferens anastomosis, vas deferens obstruction, reproductive tract injury and testicular injury. 2. Immune dysfunction; the possible cause is a decrease in the amount or activity of suppressor T lymphocytes. Under normal circumstances, a small amount of sperm antigens leaked from the blood testis and its vas deferens can activate suppressor T cells, making the process of mature B cells recognizing antigens slow, reducing the body's humoral immune response to sperm antigens, and forming immune tolerance. When the number or activity of suppressor T cells decreases and the factors that activate suppressor T cells in semen are lacking, anti-sperm antibodies can also be produced. 3. Abnormalities in seminal plasma immunosuppressive substances: Seminal plasma immunosuppressive substances can block sperm antigens, inhibit local immune responses in the reproductive tract, and have a physiological protective effect on sperm and reproductive activity. When the quality and quantity of seminal plasma immunosuppressive substances change or become abnormal, this balance can be disrupted. If the husband's semen lacks seminal plasma immunosuppressive substances, the spouse can produce anti-sperm antibodies.

The anti-sperm antibodies produced will interfere with reproduction, and the main mechanisms are: 1. Affecting sperm movement; anti-sperm antibodies bind to sperm, causing sperm agglutination, resulting in decreased sperm motility, movement obstruction, and reduced sperm survival rate. At the same time, after the sperm is bound by anti-sperm antibodies, its ability to penetrate cervical mucus is significantly inhibited. Therefore, anti-sperm antibodies can reduce the chance of pregnancy by reducing sperm motility, survival rate and inhibiting sperm from passing through cervical mucus. 2. Affecting sperm capacitation and acrosome reaction; anti-sperm antibodies can affect the activity of sperm membranes, thereby hindering or delaying capacitation. Anti-sperm antibodies can hinder sperm from releasing hyaluronidase, thereby inhibiting sperm acrosome reaction. 3. Affecting sperm passing through the zona pellucida and sperm-egg fusion; anti-sperm antibodies inhibit sperm acrosome reaction, thereby inhibiting sperm from dispersing the corona radiata and penetrating the zona pellucida, reducing sperm-egg fusion. 4. Impact on fertilized eggs; anti-sperm antibodies can interfere with the implantation of fertilized eggs, resulting in embryo absorption or miscarriage. The possible reason is that the early embryo can temporarily express various antigens during its development, some of which have cross-immunity with sperm proteins and teratomas. There are many methods for detecting anti-sperm antibodies, but there are also various shortcomings. An ideal method for detecting antibodies should meet the following conditions: 1. High sensitivity, strong specificity, and good repeatability. 2. Ability to determine the type of antibody. 3. Ability to determine the binding site of the antibody. 4. Ability to quantitatively or semi-quantitatively detect anti-sperm antibodies. The antibody titer and the binding site of the antibody on the sperm determine the extent of its damage to fertility. Anti-sperm antibodies bound to the sperm head cause greater damage to fertility, while antibodies bound to the sperm tail have less effect on fertility. Therefore, detecting antibodies on the surface of the sperm head is more practical. Anti-sperm antibody detection methods include mixed antiglobulin reaction test (MARtest), gelatin agglutination test (GAT), tissue compatibility plate agglutination test (TAT), immunobead test (IBT) and enzyme-linked immunosorbent assay (ELISA).

Alternative Names

Spermatozoon
Spermatozoa
Sperm cell
Male gamete

References

1. A S V, Dhama K, et al. Role of Antisperm Antibodies in Infertility, Pregnancy, and Potential forContraceptive and Antifertility Vaccine Designs: Research Progress and Pioneering Vision. Vaccines (Basel) l. 2019, 7(3):116.
2. Silva AF, et al. The impact of antisperm antibodies on human male reproductive function: an update. Reproduction. 2021, 162(4):R55-R71.

Documents

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QA & Reviews

Customer Reviews

References

In vitro Development of Preimplantation Caprine Embryo using Cryopreserved Black Bengal Buck Semen

INDIAN JOURNAL OF ANIMAL RESEARCH

Authors: Kumar, Rohit; Chandra, P.; Konyak, P.; Karunakaran, M.; Santra, A.; Das, Subrata K.

Abstract

The aim of the present study was to produce goat embryo in different culture media through in vitro fertilization using cryopreserved black Bengal buck semen. Total 1265 fresh cumulus oocyte complexes (COGS) were collected by aspiration method with a 19 gauge hypodermic needle, washed 5-6 times and cultured in maturation media maintaining 5% CO, level at 38.5 degrees C with maximum humidity in a incubator. After 27 h of incubation cumulus cells were stripped off from matured oocytes and transferred to acidified Tyrode's medium for zona thinning and co-incubated with in vitro capacitated sperms for fertilization in Fert-BO media. In the experiment I, fresh buck semen and in experiment II, frozen buck semen was used for in vitro fertilization after in vitro processing. After 5 h of coincubation, presumptive zygotes were washed and co-incubated with oviductal cells in three different culture media (RVCL, mSOF, KSOM) for further development. In fresh group cleavage (%) were 37.76 +/- 2.98, 39.60 +/- 1.75, 29.01 +/- 1.74, and morula formation (%) were 7.72 +/- 3.38, 6.03 +/- 1.29 and 3.00 +/- 3.00 in RVCL, mSOF and KSOM media respectively. However, in frozen group cleavage (%) were 29.17 +/- 2.56, 27.70 +/- 2.31, 24.17 +/- 1.44 in RVCL, mSOF and KSOM media respectively and morula formation (%) was 2.93 +/- 0.97 only in RVCL media. These results indicate that cryopreserved black Bengal buck semen have competence to produce embryos and could be used for embryo development in RVCL media through in vitro fertilization.

Seminal cell-free DNA and sperm characteristic's: An added biomarker for male infertility investigation

ANDROLOGIA

Authors: Di Pizio, Pierre; Celton, Noemie; Menoud, Pierre Alain; Belloc, Stephanie; Bacrie, Martine Cohen; Belhadri-Mansouri, Naima; Rives, Nathalie; Cabry, Rosalie; Benkhalifa, Moncef

Abstract

Cell-free DNA (Cf-DNA) fragments may constitute an easy-to-measure molecular tool for guiding the choice of care provided to infertile couples who benefit assisted reproductive technology (ART) programmes. Data on Cf-DNA levels in the seminal plasma of men with sperm alterations are scarce. The objective of the present study was to quantify the presence of Cf-DNA in semen by using a quantitative real-time PCR. We compared men with abnormal sperm characteristics (n = 21) with normospermic controls (n = 21). The PCR assay evidenced significantly higher mean Cf-DNA levels in patients with sperm abnormalities than in controls (2.09 versus 1.18 mu g/ml, respectively;p = .0003). The Cf-DNA levels were notably higher in men with azoospermia (3.65 mu g/ml, versus 1.34 mu g/ml in matched controls;p = .03) and men with teratozoospermia (1.80 mu g/ml, versus 1.29 mu g/ml in matched controls;p = .008). Our data report a significant association between elevated Cf-DNA levels and sperm abnormalities. These results may open up new diagnostic and prognostic perspectives in male infertility.