Bijsluiter spermmar

SpermMar IgA Test
This can be done by means of the 10 microlitres capillary immunoglobulin class of antispermatozoal antibodies in serum. Am J Reprod Immunol Microbiol, 1985, 7 : 143-147. Test for determination of Sperm Antibodies Note: To use the microcapillary pipettes, insert the end of 4. FRIBERG J: Immunoglobulin concentration in serum and seminal fluid from men with and without sperm-agglutinating antibodies. Obstet Gynecol, 1980, the pipette marked with a heavy black line into the rubber A qualitive Latex Test for detection of Sperm Antibodies of bulb (approximately 5 mm). Allow pipette to fill by capillary 5. HINTING A, VERMEULEN L and COMHAIRE F: The indirect mixed antiglobulin action to the first mark (10 microlitres). Do not draw liquid reaction test using a commercially available kit for the detection of antisperm into the bulb. Place tip of finger over the top of the bulb. antibodies in serum. Fertil Steril, 1988, 49 : 1039-1044. Holding the bulb between thumb and middle finger, gently 6. JAGER S, KREMER J and VAN SLOCHTEREN-DRAAISMA T: A simple method Store at 2° to 8°C. DO NOT FREEZE.
squeeze the bulb to expel the liquid from the pipette. of screening for antisperm antibodies in the human male. Detection of sperma- In vitro Diagnostic Reagent for Professional Use only.
tozoal surface IgG with the direct mixed antiglobulin reaction carried out on 3. Mix the sample and the Latex reagent 5 times with the untreated fresh human semen. Int J Fertil, 1978, 23 : 12-21. 7. JAGER S, KREMER J , KUIKEN J and VAN SLOCHTEREN-DRAAISMA T: 4. The cover glass is put on the mixture and the mixture is Immunoglobulin class of antispermatozoal antibodies from infertile men and observed under a light microscope using a 400x to 600x inhibition of in vitro sperm penetration into cervical mucus. Int J Androl, 1980, KIT CONTENTS
magnification. The use of phase contrast or dark field illumination may facilitate reading the slide.
8. KREMER J and JAGER S: The significance of antisperm antibodies for sperm- 1 vial SpermMar IgA Latex Particles coated with specific 5. Read the result after 3 minutes. Observe for Latex particles cervical mucus interaction. Hum Reprod, 1992, 7 : 781-784. 9. MEINERTZ H and HJORT T: Detection of autoimmunity to sperm: mixed attached to the motile sperm. Count 100 sperm to deter- antiglobulin reaction (MAR) test or sperm agglutination? A study on 537 men mine the percentage of reactive sperm. Read again after from infertile couples. Fertil Steril, 1986, 46 : 86-91. SUMMARY AND EXPLANATION
10. PARSLOW JM, POULTON TA, BESSER GM and HENDRY WF: The clinical rel- Note: Keep the preparation in a damp chamber (e.g. a petri evance of classes of immunoglobulins on spermatozoa from infertile and The presence of sperm antibodies reacting with antigen(s) on dish containing a moistened piece of filter paper). vasectomized males. Fertil Steril, 1985, 43 : 621. the spermatozoa is considered as typical and specific for 6. The diagnosis of immunological infertility is suspected 11. RUMKE P and HELLINGA: Autoantibodies against spermatozoa in sterile immunological infertility (1,4,11,15). These antibodies are men. Am J Clin Pathol, 1959, 32 : 357-363. when 10-39% of the motile spermatozoa are attached to found in approximately 8% of infertile men (17). Antisperm 12. STEDRONSKA J and HENDRY WF: The value of the mixed antiglobulin reac- latex particles; if 40% or more of the spermatozoa are tion (MAR-Test) as an addition to routine seminal analysis in the evaluation of antibodies belong to different immunological classes, but only attached, immunological infertility is highly probable.
the subfertile male. Am J Reprod Immunol, 1983, 3 : 89-91. those of the IgG and the IgA class are clinically relevant (3).
13. UELING DT: Secretory IgA in seminal fluid. Fertil Steril, 1971, 22 : 769-773. The former display cytotoxic effects and are adequately 14. VERMEULEN L and COMHAIRE F: Le test “MAR” aux particles de Latex, et detected on spermatozoa or in serum with the SpermMar IgG INDIRECT SPERMMAR TEST FOR THE DETECTION OF le test spermatoxique selon Suominen : simplification et nouveauté dans l’arse- test. Antisperm antibodies of the IgA class, which mainly have nal du diagnostic immunologique. Contraception-Fertilité-Sexualité, 1983, 11 agglutinating properties (8), rarely occur without antibodies of 15. WILSON L: Sperm agglutinins in human semen and blood. Proc Soc Exp Biol the IgG class (7), but their meaning for male infertility may be more important. Indeed, patients combining sperm antibodies 1. Allow the reagent and seminal plasma or cervical mucus 16. WORLD HEALH ORGANIZATION: Laboratory manual for the examination of of the IgA class with IgG antibodies, or presenting IgA anti- specimens to come to room temperature.
human semen and semen-cervical mucus interaction. Cambridge University bodies alone have very little chance of impregnating their 2. Inactivate the specimens by heating them at 56°C for partner through natural ways (7,9). Hence, detection of anti- 17. WORLD HEALTH ORGANIZATION: Towards more objectivity in diagnosis and bodies of the IgA class is of the utmost importance both for management of male infertility. Int J Androl, 1987, Suppl 7. 3. Dilute inactivated specimens 1/4 with Earle’s medium.
18. ZAVOS PM: Seminal parameters of ejaculate collected from oligospermic 4. Wash the motile donor sperm by letting them swim up and normospermic patients via masturbation and at intercourse with the use of The bulk of the IgA class antisperm antibodies are secreted by Silastic seminal fluid collection device. Fertil Steril, 1985, 44 : 517-520. the accessory sex glands (13). They are present on the sper- Afterwards adjust the sperm concentration to 20x106 matozoa and sometimes in seminal plasma, but usually are spermatozoa/ml with Earle’s medium.
absent in serum. Therefore, testing for antisperm antibodies of 5. Incubate 100 microlitres of the suspension of motile donor the IgA class on serum is not recommended (3). It may be con- SpermMar IgA is produced by FertiPro N.V.
spermatozoa with 100 microlitres of inactivated seminal sidered to search for sperm antibodies of the IgA class in Industriepark Noord 32, 8730 Beernem, Belgium plasma or cervical mucus, which has been diluted 1/4 in seminal plasma, in cases with low sperm concentration or oocyte culture medium, for 1 hour at 37°C.
motility, although the possible clinical meaning of these anti- 6. Add 2 ml of Earle’s medium, mix well and centrifuge for The direct SpermMar test is a test for the detection of sperm 7. Decant or aspirate the supernatant. Resuspend the pellet coating antibodies, performed on either fresh spermatozoa, or with 50 microlitres of Earle’s medium.
spermatozoa which are isolated from seminal plasma by one cycle of suspension, centrifugation and resuspension in medium. These spermatozoa are mixed with Latex particles 10 microlitres of SpermMar Latex particles which are coated with antihuman anti-IgA. The formation of This can be done by means of the 10 microlitres capillary mixed agglutination of motile spermatozoa with latex particles indicates the presence of IgA antisperm antibodies on the 9. Mix the sample and the Latex reagent 5 times with the In the indirect SpermMar test, washed motile donor sperma- 10.The cover glass is put on the mixture and the mixture is tozoa are incubated with diluted seminal plasma or cervical observed under a light microscope using a 400x to 600x mucus. If the tested fluid contains antisperm antibodies, these magnification. The use of phase contrast or dark field will bind to the donor spermatozoa and cover them. Next the illumination may facilitate reading the slide.
antibodies will be detected by means of the mixed antiglo- 11.Read the result as described for the direct SpermMar test.
buline reaction test. Performance of the indirect IgA SpermMar Note: Keep the preparation in a damp chamber (e.g. a petri test is to be considered experimental and the clinical meaning dish containing a moistened piece of filter paper). of its results has not been established.
12.For indirect testing on cervical mucus the diagnosis of immunological infertility is highly probable if 10% or more REAGENTS
of the motile spermatozoa have latex particles attached.
One reagent is supplied in each package of SpermMar IgA test : INTERPRETATION OF RESULTS
SpermMar Latex particles is a suspension of polystyrene latex particles of approximately 2.0 µm in diameter coated with When the test is performed properly, the absence of antisperm monoclonal antihuman anti-IgA serum. Mix well before use.
antibodies will be shown by freely moving spermatozoa uncovered by Latex particles. The Latex particles may, but The reagent is preserved with sodium azide at a final concen- usually do not agglutinate among themselves. In the presence of antisperm antibodies the spermatozoa will react with the particles and one, later a bunch of particles will attach to all or REAGENT PREPARATION
a proportion of the motile spermatozoa. The percentage of SpermMar
motile spermatozoa showing this mixed agglutination is The SpermMar Latex particles are ready to use, however, they directly related with the severity of the immunological reac- should be thoroughly mixed before use to provide a homoge- tion. In general, the proportion of motile spermatozoa reacting in the SpermMar IgA test is smaller than that reacting in the SpermMar IgG test, but the contrary may occasionally occur REAGENT STORAGE
(16). In rare cases there is a positive reaction in the SpermMar IgA test in the absence of any reaction in the SpermMar IgG When stored properly, SpermMar reagent is stable for 12 test, indicating the presence of antibodies of the IgA class months from date of manufacturing. SpermMar reagent must without antibodies of the IgG class. Occurence of mixed agglu- be stored at between 2° and 8°C when not in use.
tination reaction of 40% or more in semen and 10% or more in cervical mucus indicate a positive reaction to the SpermMar SPECIMEN COLLECTION AND PREPARATION
LIMITATIONS OF THE METHOD
Semen collection by masturbation is preferred. Where parti- cular circumstances discourage collection by masturbation, The direct SpermMar test can only be performed if motile specific plastic condoms are available for semen collection spermatozoa are present in the semen sample. Samples with (Male Factor Pak). Ordinary condoms cannot be used for very low sperm concentration or motility may yield false semen collection because they may interfere with the motility negative results. Performance of the indirect SpermMar IgA and viability of the spermatozoa. Ideally, semen should be test is to be considered experimental and the clinical meaning examined within 1 hour after ejaculation. Cervical mucus can of its result has not been established.
be aspirated using a device specially designed for this pur- pose (e.g. Spirette, Mucat or Aspylaire).
PERFORMANCE CHARACTERISTICS
WARNINGS AND PRECAUTIONS
All human, organic material should be considered potentially Several hundreds of semen samples have been tested with the direct mixed antiglobuline reaction and the direct Handle all specimens as if capable of transmitting HIV or SpermMar test for IgG. The results were similar in 97% of the hepatitis. Always wear protective clothing when handling cases. In 3% of the cases the SpermMar test detected anti- bodies whereas the mixed antiglobulin reaction test using SpermMar IgA contains 0.1% Bovine Serum Albumin of US coated red blood cells was negative. In such cases the pro- portion of spermatozoa reacting in the SpermMar test usually was low (8). The results of the SpermMar IgA test were proven PROCEDURE
accurate after comparison with immunofluorescence and Materials provided: (included in the complete kit only)* Since sperm antibodies of the IgA class are mostly secreted by Microcapillary pipettes calibrated at 10 microlitres* the accessory glands, they can be present on spermatozoa but not in serum. However, the majority of these patients do have sperm antibodies of the IgG class in their serum. Testing for antisperm antibodies of the IgA class in seminal plasma or Light microscope (400x to 600x magnification) cervical mucus is to be considered experimental and the Earle’s medium (for indirect SpermMar test only) clinical meaning of its results has not been established.
DIRECTIONS FOR USE
BIBLIOGRAPHY
DIRECT SPERMMAR TEST FOR THE DETECTION OF 1. BOETTCHER B, HJORT T, RUMKE P, SHULMAN S and VYAZOV (eds): Auto- and isoantibodies to antigens of the human reproductive system. 1. Results of an international comparative study of antibodies to spermatozoa and other anti- gens detected in sera from infertile patients deposited in the WHO reference bank for reproductive immunology. Acta Pathol Microbiol Scand, 1977, 258 1. Allow reagents and specimens to come to room temperature.
2. BRONSON R, COOPER G and ROSENFELD D: Sperm antibodies: their role in infertility. Fertil Steril, 1984, 42 : 171-183. 10 microlitres of SpermMar Latex particles 3. CLARKE GN, STOJANOFF A, CAUCHI MN and JOHNSTON WIH: The SpermMar IgG Test
inhibition of in vitro sperm penetration into cervical mucus, IntJ. Androl., HIV or hepatitis. Always wear protective clothing when Direct and Indirect Test for Determination 1. Allow all reagents and specimens to come to room 7. RUMKE P.: The origin of immunoglobulins in semen. Clin. Exp. Immunol., Although SpermMar IgG Positive and Negative controls have been tested for HIV and hepatitis the user should always wear 8. SHULMAN J.F. and SHULMAN S.: Methylprednisolone treatment of 2. Inactivate the serum specimens by heating them at 56°C protective clothing when handling the control sera.
immunologic infertility in the male. Fertil Steril, 1982, 38 : 591-599. A qualitative Latex Test for detection of Sperm Antibodies.
for 30 minutes if glass test-tubes are used, 45 minutes if 9. STEDRONSKA J. and HENDRY W.F.: The value of the mixed antiglobulin reaction (MAR-Test) as an addition to routine seminal analysis in the evalu- PROCEDURE
3. Adjust the pH (by adding 0.1N NaOH or HCl) of Earle’s or ation of the subfertile couple. Am. J. Reprod. Immunol., 1983, 3 : 89-91. Store at 2° to 8° C - Do Not Freeze.
other medium* with similar composition to 7.4 - 7.5.
10. VERMEULEN L. and COMHAIRE F.H.: Le test <MAR> aux particules de Latex, et le test spermatoxique selon Suominen : simplification et nouveauté In Vitro Diagnostic Reagent for Professional Use Only.
4. Wash the motile donor spermatozoa by letting them swim dans l’arsenal du diagnostique immunologique. Contraception-Fertilité- up in the pH adjusted medium (pH = 7.4 - 7.5). Swim up or
Sexualité, 1983, 11 (suppl.) : 381-384. has to be done in 5 ml glass or sterile plastic test-tubes 11. W.H.O. 1984: Workshop on the standardized investigation of the infertile with round bottom at 37°C for 45 minutes.
couple, moderator P. Rowe, coordinator M. Darling. Fertil Steril (eds) R.F. KIT CONTENTS
Adjust the sperm concentration to 2x107 sp/ml with HARRISON, J. BONNAR and W. THOMPSON. Publ. M.T.P.-Press Ltd. (Lancaster, Boston, The Hague, Dordrecht), 1984, 427-431. 1 Vial SpermMar Latex Particles 0.7 ml.
5. Serially dilute inactivated serum specimen 1/16 with 12. W.H.O.: Laboratory manual for semen analysis and sperm cervical Light Microscoop (400x or 600x magnification) mucus interaction. Publ. Cambridge University Press, 1987. medium (pH = 7.4 - 7.5) in a titer plate.
13. COMHAIRE F.H., HINTING A., VERMEULEN L., SCHOONJANS F. and 6. Mix 50 microlitres of diluted inactivated serum specimen GOETHALS I.: Evaluation of the direct and indirect mixed antiglobuline reac- SUMMARY AND EXPLANATION
(step 5) with 50 microlitres of the washed motile donor tion with latex particles for the diagnosis of immunological infertility. Int. J. DIRECTIONS FOR USE
sperm (step 4) in a free well on the titer plate. Let incubate The presence of sperm antibodies reacting with antigens on 14. KREMER J. and JAGER S.: The significance of antisperm antibodies for 1. Allow all reagents and specimens to come to room the spermatozoa is considered as typical and specific for sperm-cercical mucus interaction. Hum. Reprod., 1992, 7 : 781-784. 15. KAY D.J. and BOETTCHER B.: Comparison of the SpermMar test with immunological infertility (2,4,11). Sperm antibodies belong to 10 microlitres of the sperm-serum mixture 2. Wash the motile donor spermatozoa by letting them swim currently accepted procedures for detecting human sperm antibodies. two immunological classes; IgA and IgG antibodies. There are 10 microlitres of SpermMar Latex Particles up in the pH adjusted Earle's* medium (pH = 7.4 - 7.5).
Reprod. Fer. Dev., 1992, 4, 175-181. some data indicating IgA to be more clinically important than Swim up has to be done in 5 ml glass or sterile plastic 16. ACKERMAN S., Mc GUIRE G., FULGHAM D.L. and ALEXANDER N.: An IgG antibodies. However, IgA antibodies rarely occur without 8. Mix the sample and the Latex reagent 5 times with the evaluation of a commercially available assay for the detection of antisperm test-tubes with round bottom at 37°C for 45 minutes.
IgG antibodies. Therefore, testing for IgG antibodies is suffi- antibodies. Fertil Steril 1988, 49 : 732-734. Adjust the sperm concentration to 2x107 sp/ml with cient as a routine screening method (6,7,14).
9. Also mix the Antiserum with the Latex reagent and sample The direct SpermMar Test is performed by mixing fresh, 3. Mix 50 microlitres of control serum with 50 microlitres of untreated semen with latex particles that have been coated 10.The cover glass is put on the mixture and the mixture is the washed motile donor sperm in a free well on the titer with human IgG. To this mixture a monospecific antihuman IgG observed under a light microscope using a 400x or 600x SpermMar IgG is produced by FertiPro N.V.
plate. Let incubate for 60 minutes at 37°C.
antiserum is added. The formation of agglutinates between magnification (phase contrast or dark field illumination Industriepark Noord 32, 8730 Beernem, Belgium.
particles and motile spermatozoa indicates the presence of may also be used to facilitate reading).
10 microlitres of the sperm-serum mixture IgG antibodies on the spermatozoa (1,5,9,10). In the Indirect 11.Read the results after 2-3 minutes. Observe for latex par- 10 microlitres of SpermMar Latex Particles SpermMar Test washed motile donor spermatozoa are incu- ticles attached to motile sperm. Count 100 spermatozoa to bated with diluted decomplemented patient serum of male or dertermine the percentage reactive sperm. If no attach- 5. Mix the sample and the Latex reagent 5 times with the female origin. If the serum contains antisperm antibodies, ment of particles to sperm is observed, read again after these will cover the donor spermatozoa which will react posi- 6. Also mix the Antiserum with the Latex reagent and sample tively in a subsequent SpermMar Test.
Note: Keep the preparation in a damp chamber (e.g. a petri dish containing a moistened piece of filter paper). 7. The cover glass is put on the mixture and the mixture is REAGENTS
observed under a light microscope using a 400x or 600x * Any oocyte incubation medium may be used, provided it magnification (phase contrast or dark field illumination Two reagents are supplied in each package of SpermMar Test: does not contain serum or albumin. We recommend either may also be used to facilitate reading).
Earle’s, Mendosa’s, HAM’s F-10 or Ringer’s lactate medium.
8. Read the results after 2-3 minutes. Observe for latex par 1. SpermMar Latex Particles is a suspension of polystyrene ticles attached to motile sperm. Count 100 spermatozoa to latex particles of approximately 2.0 micron in diameter Use medium without HCO3. If medium with low HCO3 dertermine the percentage reactive sperm. If no attach- coated with human IgG. Mix well before use. Volume: 0.7 ml.
concentration is used, incubation should take place in a ment of particles to sperm is observed, read again after 10 2. SpermMar antiserum: monospecific antiserum directed 5% CO2 incubator or after adding HEPES buffer (9 ml towards the Fc fragment of human IgG. Volume: 0.7 ml.
Note: Keep the preparation in a damp chamber (e.g. a petri dish containing a moistened piece of filter paper). The two reagents are preserved with sodium azide at a final To prevent evaporation during incubation, always cover * Any oocyte incubation medium may be used, provided it does not contain serum or albumin. We recommend either REAGENT PREPARATION
Earle’s, Mendosa’s, HAM’s F-10 or Ringer’s lactate medium.
SpermMar Latex Particles are ready to use, however, they When the test is properly performed, the absence of sperm Use medium without HCO3. If medium with low HCO3 should be thoroughly mixed before use to provide a homoge- antibodies will be shown by freely moving spermatozoa concentration is used, incubation should take place in a neous suspension. SpermMar Antiserum is ready to use.
uncovered by latex particles. The latex particles themselves 5% CO2 incubator or after adding HEPES buffer (9 ml will form growing agglutinates thus proving the reactivity of REAGENT STORAGE
SpermMar IgG Test
the reagents. In the presence of sperm antibodies however, the spermatozoa will be partially covered by latex particles. In To prevent evaporation during incubation, always cover positive and negative controls
When stored properly, SpermMar reagents are stable for 18 some cases the spermatozoa might even be immobilized by Positive and negative controls for use with months from the date of manufacturing. SpermMar reagents the massive amount of adherent latex particles. In the direct must be stored at 2° to 8°C when not in use. DO NOT FREEZE.
SpermMar Test, the diagnosis of immunological infertility is the Indirect Test for Determination of Sperm suspected when 10-39% of the motile spermatozoa are SPECIMEN COLLECTION AND PREPARATION
covered by latex particles; if 40% or more of the spermatozoa When the test is properly performed, the absence of sperm are covered, immunological infertility is highly probable.
antibodies will be shown by freely moving spermatozoa The semen should be collected by masturbation or by other Additional tests should confirm the diagnosis. Whenever a uncovered by latex particles. The latex particles themselves methods recommended by the physician. Preferentially, positive result is obtained it is recommended to perform the will form growing agglutinates thus proving the reactivity of semen should be examined within 1 hour after ejaculation.
SpermMar IgA test. In the indirect SpermMar Test, the occur- In Vitro Diagnostic Reagent for Professional Use Only.
the reagents. In the presence of sperm antibodies however, Serum is obtained by venous puncture.
rence of 40% or more reaction between the coated latex the spermatozoa will be partially covered by latex particles. In particles and motile spermatozoa is the lower limit of some cases the spermatozoa might even be immobilized by WARNINGS AND PRECAUTIONS
the massive amount of adherent latex particles.
PRODUCT CODES AND KIT CONTENTS
The SpermMar IgG positive control test should yield 80% or All human, organic material should be considered potentially LIMITATIONS OF THE PROCEDURE
more of the motile spermatozoa covered with latex particles.
infectious. Handle all specimens as if capable of transmitting SPMG_P: 1 vial with 2.5 ml of positive control serum for the The SpermMar IgG negative control should yield 20% or less HIV or hepatitis. Always wear protective clothing when hand- The direct SpermMar-Test can only be performed if motile spermatozoa covered with latex particles.
spermatozoa are present in the semen. Samples with poor SPMG_N: 1 vial with 2.5 ml of negative control serum for the SpermMar IgG latex particels contains 0.1% Bovine Serum motility may yield false negative results.
LIMITATIONS OF THE PROCEDURE
SpermMar IgG latex particels are coated with human IgG, all PERFORMANCE CHARACTERISTICS
SUMMARY AND EXPLANATION
The indirect SpermMar-Test can only be performed if motile materials used have been tested by their original manufacturer spermatozoa are present in the semen. Samples with poor for Hepatitis B, Hepatitis C and HIV.
The presence of sperm antibodies reacting with antigens on motility may yield false negative results.
the spermatozoa is considered as typical and specific for PROCEDURE
Several hundreds of semen samples were tested with the immunological infertility (1,2,5). Sperm antibodies belong to PERFORMANCE CHARACTERISTICS
direct MAR-Test (mixed antiglobulin reaction based on red two immunological classes; IgA and IgG antibodies. There are Materials provided: (Included in complete kit only)* blood cells) and with the SpermMar-Test. The results were some data indicating IgA to be more clinically important than Using the value of 40% reaction between motile spermatozoa similar in 97% of the cases. In 3% of the cases the MAR-Test IgG antibodies. However, IgA antibodies rarely occur without and coated Latex particles as the lower limit of significant based on red blood cells was negative while the SpermMar- IgG antibodies. Therefore, testing for IgG antibodies is activity, the indirect SpermMar-Test was found positive in Test detected antibody coated spermatozoa, though in sufficient as a routine screening method (3,4,6).
some cases with negative results of the Tray Agglutination relatively small numbers (<40%), thus proving the higher In the Indirect SpermMar Test washed motile donor spermatozoa Test, or other currently accepted procedures.
Microcapillary pipettes calibrated at 10 microlitres* sensitivity of the SpermMar Test (10,16).
are incubated with diluted, decomplemented patient serum of The SpermMar-Test was proven easier to use and more male or female origin. If the serum contains antisperm anti- sensitive (7). A false negative indirect SpermMar-Test in com- bodies, these will cover the donor spermatozoa which will parison with the Tray Agglutination Test occured in cases with Required supplementary materials: (Both Kits) react positively in a subsequent SpermMar Test.
IgM in serum, the clinical significance of which is doubtful. It Light Microscoop (400x or 600x magnification) Using the value of 40% reaction between motile spermatozoa The SpermMar IgG positive control contains ready-to-use is recommended to confirm a positive result of the indirect Earle’s Medium (for Indirect SpermMar-Test only) and coated Latex particles as the lower limit of significant patient serum with antisperm antibodies levels higher than SpermMar-Test by additional tests for the detection of agglu- activity, the indirect SpermMar-Test was found positive in tinating activity (Tray Agglutination Test) and of cytotoxic DIRECTIONS FOR USE
some cases with negative results of the Tray Agglutination The SpermMar IgG negative control contains ready-to-use activity, such as the ATP-Release cytotoxicity test. The latter Test, or other currently accepted procedures.
patient serum with antisperm antibodies levels of below 20%.
tests will also assess the type of immunological effect The SpermMar-Test was proven easier to use and more exerted by the antisperm antibodies.
sensitive (15). A false negative indirect SpermMar-Test in REAGENTS
1. Allow the reagents and specimens to come to room comparison with the Tray Agglutination Test occured in cases BIBLIOGRAPHY
with IgM in serum, the clinical significance of which is doubt- Each package of SpermMar IgG Positive and Negative control ful. It is recommended to confirm a positive result of the 1. COMHAIRE F.H. and KUNNEN M.: Factors affecting the probability of concep- tion after treatment of subfertile men with varicocele by transcatheter emboli- indirect SpermMar-Test by additional tests for the detection of Decomplemented patient serum diluted in FertiCult Flushing sation with Bucrylate. Fertil Steril, 1985, 43 : 781-786. 10 microlitres of SpermMar Latex Particles agglutinating activity (Tray Agglutination Test) and of cytotoxic medium without human serum albumin. Volume: 2.5 ml 2. GOLOMB J., VARDINON H., HOMONNAI Z.T., BRAF Z. and YUST I.: activity, such as the ATP-Release cytotoxicity test. The latter Sodium azide has been added to this solution at a final con- Demonstration of antispermatozoal antibodies in varicocele-related infertility This can be done by means of the provided 10 microlitres tests will also assess the type of immunological effect with an enzym-linked Immunosorbent assay (ELISA). Fertil Steril, 1986, 45 : exerted by the antisperm antibodies.
Note: To use the microcapillary pipettes: Insert the end of the 3. JAGER S., KREMER J., KUIKEN J. and VAN SLOCHTEREN-DRAAISMA T.: REAGENT PREPARATION
pipette marked with the heavy black line into the rubber bulb Immunoglobulin class of antispermatozoal antibodies from infertile men and BIBLIOGRAPHY
inhibition of in vitro sperm penetration into cervical mucus, IntJ. Androl., 1980, (approximately 5 mm). Allow the pipette to fill by capillary SpermMar IgG Positive and Negative control are ready to use.
action to the first mark (10 microlitres). Do not draw liquid 1. BOETTCHER B., HJORT T., RUMKE Ph., SHULMAN S. and VYAZOV O.E. Allow to warm to room temperature before use.
4. RUMKE P.: The origin of immunoglobulins in semen. Clin. Exp. Immunol., into the rubber bulb. Holding the bulb between the thumb and (eds.): Auto and isoantibodies to antigens of the human reproductive system. 1. Results of an international comparitive study of antibodies to spermato- the middle finger, gently squeeze the bulb to expel the liquid REAGENT STORAGE
5. W.H.O. 1984: Workshop on the standardized investigation of the infertile zoa and other antigens detected in sera from infertile patients deposited in couple, moderator P. Rowe, coordinator M. Darling. Fertil Steril (eds) R.F. the WHO Reference bank for reproductive immunology. Acta Pathol Microbiol 3. Mix the sample and the Latex reagent 5 times with the HARRISON, J. BONNAR and W. THOMPSON. Publ. M.T.P.-Press Ltd. (Lancaster, When stored properly, SpermMar IgG control sera are stable Boston, The Hague, Dordrecht), 1984, 427-431. 2. COMHAIRE F.H. and KUNNEN M.: Factors affecting the probability of con- for 18 months from the date of manufacturing. SpermMar IgG 6. KREMER J. and JAGER S.: The significance of antisperm antibodies for 4. Also mix the Antiserum with the Latex reagent and sample ception after treatment of subfertile men with varicocele by transcatheter control sera must be stored at 2° to 8°C when not in use.
sperm-cercical mucus interaction. Hum. Reprod., 1992, 7 : 781-784. embolisation with Bucrylate. Fertil Steril, 1985, 43 : 781-786. 7. KAY D.J. and BOETTCHER B.: Comparison of the SpermMar test with 5. The cover glass is put on the mixture and the mixture is 3. FRIBERG J.: Immunoglobulin concentration in serum and seminal fluid SPECIMEN COLLECTION AND PREPARATION
currently accepted procedures for detecting human sperm antibodies. Reprod. from men with and without sperm-agglutinating antibodies. Obstet. observed under a light microscope using a 400x or a 600x magnification (phase contrast or dark field illumination 4. GOLOMB J., VARDINON H., HOMONNAI Z.T., BRAF Z. and YUST I.: The donor semen should be collected by masturbation or by may facilitate reading of the slides).
Demonstration of antispermatozoal antibodies in varicocele-related infertil- other methods recommended by the physician. Preferentially, 6. Read the result after 2-3 minutes. Observe for latex par- ity with an enzym-linked Immunosorbent assay (ELISA). Fertil Steril, 1986, semen should be examined within 1 hour after ejaculation.
ticles attached to motile sperm. Count 100 spermatozoa to SpermMar IgG is produced by FertiPro N.V.
Serum is obtained by venous puncture.
dertermine the percentage reactive sperm. If no attach- 5. JAGER S., KREMER J. and VAN SLOCHTEREN-DRAAISMA T.: A simple Industriepark Noord 32, 8730 Beernem, Belgium.
method of screening for antisperm antibodies in the human male. Detection ment of beads to sperm is observed, read again after WARNINGS AND PRECAUTIONS
of spermatozoal surface IgG with the direct mixed antiglobulin reaction car- ried out on untreated fresh human semen. Int. J. Fertil., 1978, 23 : 12-21. Note: Keep the preparation in a damp chamber (e.g. a petri 6. JAGER S., KREMER J., KUIKEN J. and VAN SLOCHTEREN-DRAAISMA T.: All human, organic material should be considered potentially dish containing a moistened piece of filter paper). Immunoglobulin class of antispermatozoal antibodies from infertile men and infectious. Handle all specimens as if capable of transmitting

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