Abnormalities for chromosomes 13 and 21 detected in spermatozoa from infertile men

Sperm samples from infertile men with oligozoospermia or teratozoospermiawere studied by multicolour fluorescence in-situ hybridization(FISH) using DNA probes for chromosomes 13 and 21. A total of90 809 sperm nuclei from nine infertile men and 182 799 spermnuclei from 18 control donors were analysed. There was a highlysignificant increase in the frequency of spermatozoa disomicfor chromosome 13 in infertile patients (0.28%) compared tocontrol donors (0.13%) (two-tailed Z statistic P <0.0001and for chromosome 21 (0.48% in infertile men versus 0.37% incontrols, P <0.0001). Also there was a significantly increasedfrequency of diploid spermatozoa in infertile men (0.85%) comparedto control donors (0.66%) (P <0.0001). Our previous studieson these same infertile patients demonstrated increased frequenciesof sperm disomy for chromosomes 1 and XY. This suggests thatinfertile men, who are prime candidates for intracytoplasmicsperm injection, may be at a very small increased risk of aneuploidoffspring.     

Lead, magnesium, selenium and zinc in human seminal fluid: comparison with semen parameters and fertility

The concentrations of lead, magnesium, selenium and zinc inseminal fluid from men with variable semen quality (sperm morphology,density and motility) and fertility were determined by atomicabsorption spectrometer without or with Zeeman background correction.The mean (?SD) concentration of selenium in the samples (n =142) was 28.8 ? 9.5 µg/l, which was about a third of thecorresponding serum value (77.8 ? 13.3 µg/l, n = 140).The serum selenium level was significantly (P < 0.001) higherin infertile than in fertile men, but the seminal fluid didnot show such a difference. No correlation was obtained betweenselenium values in seminal plasma and sperm density or motility.The levels of lead in seminal fluid were very low with no correlationto the levels of magnesium, selenium and zinc or the semen qualities.The seminal fluid lead concentration was significantly (P <0.001) higher in infertile (3.6 ? 3.2 µg/l, n = 79) thanin fertile men (1.7 ? 1.0 µg/l, n = 39). Magnesium (103.5? 49.2 mg/l, n = 90) and zinc (141.1 + 71.7 mg/l, n = 157) concentrationsin seminal fluid were comparable with previous reports. Bothminerals showed a positive correlation to the seminal fluidselenium, while only zinc displayed a borderline correlationwith sperm density. The present findings indicate that the determinationof seminal fluid selenium may not offer any advantages overzinc and magnesium measurement in the fertility assessment andits role in human semen remains obscure. The low lead concentrationsin the present material is a clear indication of low industrialexposure.     

Studies on Sperm Survival and Motility in the Presence of Cytotoxic Sperm Antibodies

ABSTRACT: The effect of cytotoxic sperm antibodies and native complement in the serum and secretions from 40 fertile and 93 infertile couples on in vitro sperm survival and motion characteristics was studied. Sperm survival in vitro was unaffected by sera from fertile and infertile subjects without cytotoxic sperm antibodies and from infertile men with antibodies to control but not to autologous sperm. Sperm survival was reduced (P < .001) by sera from infertile men with antibodies to autologous sperm or to antologous and control sperm and from women with cytotoxic antibodies to sperm from both. Sera from fertile couples without sperm antibodies enhanced sperm swimming speed and motility index (P < .0001). Sera from infertile women with or without cytotoxic sperm antibodies did not affect sperm motility. Sperm survival and motility were reduced by seminal plasma from infertile men with cytotoxic antibodies to autologous and/or control sperm. Seminal plasma from fertile men enhanced sperm survival. Cervical mucus from infertile women with antibodies to autoimmune husbands' sperm or to husbands' and control sperm inhibited sperm motion, whereas cervical mucus from infertile women without sperm antibodies and women with antibodies to control sperm failed to have any effect. It is concluded that cytotoxic sperm antibodies developed through exposure to sperm antigens in autoimmune infertile men decrease in vitro sperm survival and/or motility.     

AZFb deletions predict the absence of spermatozoa with testicular sperm extraction: preliminary report of a prognostic genetic test

Genetic abnormalities, including partial deletions of the Y-chromosome,are commonly detectable in men with non-obstructive azoospermia(NOA). NOA can be treated using testicular sperm extraction(TESE) with intracytoplasmic sperm injection (ICSI). Recentstudies have shown that the presence of deletions involvingthe AZFc region do not appear to affect the chance of retrievingspermatozoa or have a significant impact on fertilization orpregnancy rates with ICSI. We investigated the effect of Y-chromosomepartial deletions on the chance of sperm retrieval with TESE.Eighty attempts at sperm retrieval were performed using TESEon men who were previously evaluated for Y-chromosome partialdeletions. Y-chromosome analysis was performed using a polymerasechain reaction (PCR)-based technique with 35 sequence-tagged-sites.Of the 80 men, nine (11%) had partial Y-chromosome deletionsdetected. Two azoospermic men with AZFc deletions had successfulsperm retrieval, ICSI and a subsequent clinical pregnancy. Sevenmen had deletions involving the AZFb region (three men had isolatedAZFb deletions, one had AZFa, AZFb and AZFc deleted, and threehad AZFb and AZFc deleted). None of the seven men had spermatozoaextracted by TESE, a result that is significantly differentfrom the overall 64% (47/73) sperm retrieval rate achieved atour centre (P = 0.001). Two men with AZFb deletions had cellsconsistent with round spermatids identified and injected intooocytes without effecting any normal fertilizations. Althoughpreliminary, these results suggest that the presence of an AZFbdeletion is a significantly adverse prognostic finding for TESE.Men with AZFb deletions should be apprised of these resultsbefore attempting TESE-ICSI. Alternatives such as donor inseminationor adoption should be considered or therapy delayed until improvedresults with round spermatid injections are likely.     

Cystic fibrosis mutation screening in healthy men with reduced sperm quality

The majority of men with cystic fibrosis (CF) are infertiledue to a bilateral congenital absence of the vas deferens (CBAVD).However, clinically affected CF patients present a spectrumof genital phenotypes ranging from normal fertility to severelyimpaired spermatogenesis and CBAVD. Recently, it has becomeapparent that CF can manifest itself as isolated CBAVD in theabsence of other clinical symptoms. The present study was undertakento test the possible involvement of the CF gene in the aetiologyof male infertility other than CBAVD. Semen specimens from 127unrelated healthy males with various diagnoses of reduced spermquality were screened for a panel of 13 mutations in the cysticfibrosis transmembrane conductance regulator (CFTR) gene. Fourteenof 80 (17.5%) healthy men with infertility due to reduced spermquality and 3 of 21 (143%) men with azoospermia had at leastone CF mutation (one azoospermic male was a compound hetero-zygote).The frequency of mutations in our sample of infertile maleswas significantly higher than the expected CF carrier frequencyin the local population (P = 0.00139). No mutations were foundin a control group of 26 individuals with normal semen parameters.This increased frequency of CF mutations in healthy men withreduced sperm quality and in men with azoospermia without CBAVDsuggests that the CFTR protein may be involved in the processof spermatogenesis or sperm maturation apart from playing acritical role in the development of the epididymal glands andthe vas deferens.     

Deterioration of sperm quality in young healthy Belgian men

We have retrospectively analysed the sperm characteristics of416 consecutive healthy young men who presented themselves inthe past 19 years as candidate sperm donors. Ejaculate volumeincreased slightly (P = 0.067), and average sperm concentrationdecreased (P = 0.035) by 12.4xlO6ml over the observation period,so that sperm count per ejaculate remained unchanged (P = 0.91).In contrast, sperm morphology (r = –0.23, P < 0.0001),rapid progressive motility (r = –0.42, P < 0.0001)and total motility (r = –0.33, P < 0.0001) presentedan important and time-related decrease. When a quadratic modelwas used rather than a linear one to analyse the data on rapidprogressive motility, there appeared to have been no furtherdecline since 1990. The average proportion of spermatozoa withnormal morphology decreased from 39.2% in the period 1977–1980to 26.6% in 1990–1995 (P < 0.0001), and the mean percentageof spermatozoa with rapid progressive motility decreased from52.7 to 31.7% (P < 0.0001). The percentage of candidate donorswith sperm characteristics below the 5th percentile cut-offvalue of a normal fertile population increased from 13 to 54%during the observation period (P < 0.0001). Since the techniqueof semen analysis has remained essentially unchanged in-so-faras has been practically possible, as has the method of recruitmentof candidate sperm donors, the observed deterioration of spermcharacteristics is considered to reflect degeneration of spermproduction among men aged between 20 and 40 years.     

Autoantibodies and antisperm antibodies in sera and follicular fluids of infertile patients; relation to reproductive outcome after in-vitro fertilization

Immune reactions have effects at various concentrations in thereproductive process and autoantibodies may have an impact onfertility and the outcome of assisted conception. We measuredthe prevalence of and relation between antibodies to smoothmuscle, nuclear, phospholipid and sperm antigens, and concentrationsof immunoglobulins G, M and A and complement components C3 andC4, in the sera and follicular fluids of women with unexplainedinfertility (n = 30), endometriosis (n = 20), tubal infertility(n = 50) and the sera of 20 normal non-pregnant women. We assessedfertilization and successful pregnancy rates in relation toantibody status of infertile women after in vitro fertilization.All antibodies had a higher prevalence in infertile women comparedwith controls and this was significant for smooth muscle antibodyin endometriosis (P < 0.05); anticardiolipin antibody intubal infertility P < 0.05); and antisperm antibody in alltypes of infertility (P < 0.001). There was no relation betweenpresence of specific antibodies in serum or between serum andfollicular fluids. Total biochemical pregnancy rate was higherwith endometriosis (P = 0.05) but clinical pregnancy and livebirth rates did not differ between groups or in relation toantibody status. Significant differences in immunoglobulin andcomplement components occurred in women with and without successfulbiochemical pregnancy.     

Immunology: Formation of antisperm antibodies in women treated with Fallopian tube sperm perfusion

Fallopian tube sperm perfusion (FSP) is a combination of ovarianstimulation and intra-uterine insemination using a large volume(4 ml) of inseminate containing 10⁷–10⁸ spermatozoa. Theinseminate will flush the Fallopian tubes and some of it willend up in the pouch of Douglas. In the present study, we haveinvestigated whether the FSP method will result in the formationof serum antisperm antibodies in the female. A total of 184treatment cycles were given to 128 women. The indications fortreatment were: unexplained infertility (n = 35), various infertilitydiagnoses (n = 28) and donor insemination (n = 65). Prior totreatment, 11 (8.6%) women had a positive tray-agglutinationtest (Friberg) and/or a positive immunobead test. After completingone to four treatment cycles, another six (4.7%) women had developedserum antisperm antibodies. The antibodies induced by the treatmentwere of isotype IgM and directed against the tailtip of thespermatozoa. Two of the women, who prior to the treatment hadantisperm antibodies, showed an increase in antibody titre duringtreatment. There was no statistically significant differencein the pregnancy rate between the women with antisperm antibodiesand the women without. In our opinion, the small risk of developingantisperm antibodies is no contra-indication for treating infertilecouples with FSP.     

Sperm membrane protein profiles of fertile and infertile men: identification and characterization of fertility-associated sperm antigen

BACKGROUND: Male infertility is the major cause of conception failure in about 25% of all infertile couples. Understanding the causes of male infertility depends to a certain extent on the proteins present on the spermatozoa. The study aim was to investigate first, whether there is any difference in the expression of sperm membrane proteins between fertile and infertile males; and second, whether there is any functional significance of these proteins in the spermatozoa. METHODS: Six different protocols were employed to extract sperm membrane proteins. A 57 kDa protein was identified and purified using different chromatographic techniques. The homogeneity and isoelectric point of the protein was confirmed by 2D-electrophoresis. The protein was characterized by immunofluorescence, ELISA, flow cytometry, SDS-PAGE and Western blot analysis. The role of 57 kDa protein in sperm-oocyte binding was studied in vitro. RESULTS: All six sperm extracts of normozoospermic and infertile subjects showed 16-18 major and 12-15 minor protein bands. However, in one of the methods, the lysis buffer containing N-octyl-beta-D-glycopyranoside (NOG) resulted in an additional protein band at the 57 kDa region in 95% of normozoospermic samples. The protein was either absent (approximately 80%) or negligible (approximately 20%) in infertile subjects. The protein was localized to the head of non-acrosome-reacted spermatozoa (NAR), and shifted to the equatorial segment in acrosome-reacted (AR) spermatozoa. The antibody directed against purified 57 kDa protein inhibited binding of human sperm to zona-free hamster oocytes in a dose-dependent manner. CONCLUSIONS: The results suggest that lack and/or low expression of 57 kDa protein may be one of the reasons for infertility in men. Therefore, the protein could be used as a marker for sperm quality in men.     

Females' isoimmunity to sperm is associated with sperm autoimmunity in their husbands

One hundred twenty-five fertile couples and 334 infertile couples were tested for the presence of cytotoxic and hemagglutinating antibodies to sperm. Elevated titers of sperm antibodies were absent in both partners of fertile couples. Of 79 infertile males with levels of sperm antibodies in the previously established negative range, 97% had wives who also had low titers of sperm antibodies. Of 255 infertile males positive for serum hemagglutinating antibodies, 56% had wives whose serum contained significant circulating hemagglutinating antibodies, while 93 of 202 (46%) males with significant cytotoxic antibody titers had wives whose serum contained elevated cytotoxic antibody titers. The females developed elevated titers of sperm antibodies in the serum and cervical mucus if their husbands had significant titers of hemagglutinating and cytotoxic sperm antibodies in the serum and seminal plasma samples. Females' isoimmunity to sperm was significantly associated with their husbands' autoimmunity to sperm and infertility.     

Fertilization and early embrology: A sequential analysis of the effect of progesterone on specific sperm functions crucial to fertilization in vitro in infertile patients

The objective of these studies was to evaluate the modulatoryeffect(s) of progesterone on sperm functions crucial to fertilizationin infertile men with abnormal sperm parameters. A prospective,controlled study applying a sequential diagnostic analysis capableof identifying specific dysfunctions of the male gamete wasperformed. Patients (n = 14) were allocated to the study groupif they had a history of infertility of >1 year durationand after semen evaluation showed teratozoospermia (< 14%normal sperm forms as diagnosed by strict criteria) or terato-asthenozoospermia(< 50% progressive motility). After swim-up separation ofthe motile sperm fraction, the following functions were assessedwith and without previous exposure to progesterone (1.0 µg/ml):acrosome reaction (using Pisum sativum agglutinin), hyperactivatedmotility (using a computerized semen analyser), sperm-zona pellucidabinding (in the hemizona assay), sperm-zona pellucida penetration(in a sperm-zona penetration assay), and sperm-oocyte penetration(using the hamster zona-free oocyte/sperm penetration assay).Progesterone did not affect the percentage of acrosome-reactedspermatozoa after 1 or 3 h of incubation. Hyperactivated motilitywas significantly enhanced by progesterone after 1 h (12 ±4 versus 6 ± 2% in controls; P < 0.02). Although progesteronedid not affect sperm-zona binding, it significantly enhancedboth sperm-zona pellucida penetration (27 versus 12% in controls;P = 0.03) and sperm-oocyte penetration (15 versus 8% in controls;P < 0.05). Because those sperm functions enhanced by progesteroneare crucial to fertilization, the steroid may have value inthe treatment of some male-factor patients undergoing assistedreproductive therapy.     

The value of intrauterine insemination with washed husband's sperm in the treatment of infertility

Twenty-nine infertile couples were treated by intrauterine insemination(IUI)of washed sperm from a sub-fertile husband (n = 16), incases of gynaecological (n = 3), combined (n = 4) or idiopathic(n = 6) infertility; 116 treatment cycles redted in 11 ongoingpregnancies. Between 0.25 and 0.45 ml of capacitation medium,containing at least 500 000 pretreated spermatozoa, were inseminated.Pretreatment of first split fractions was performed by centrifugationand swimming up of motile spermatozoa. The pregnancy per cycleindex (P/C) for IUI was 9.5% for a total of 37.9% of all couplestreated achieving pregnancy. These results suggest a substantialbenefit compared with a calculated six months' cumulative pregnancyrate of 4.2% independent of treatment, for this infertile population.The value of IUI in selected cam of infertility seems obviousbut needs further investigation.     

White cell subpopulations in autoimmune infertile men and their wives with cytotoxic sperm antibodies

Sixty six infertile men with cytotoxic sperm antibodies had fewer and lower percentages of OKT4-positive helper/inducer T lymphocytes p = 0.004 and 0.003, respectively), than the fertile (n = 14) and infertile (n = 12) antibody-negative controls. Ratio of OKT4/OKT8 was decreased (p less than 0.01), while numbers of OKDR and BD-Leu12 positive predominantly B lymphocytes were increased (p less than 0.0001) in infertile men with sperm antibodies. T lymphocytes positive for OKT3, OKT11 and OKT4 and the OKT4/OKT8 ratio were decreased in infertile women with sperm antibodies, while B lymphocytes positive for OKDR and BD-Leu12 were increased in infertile women with or without sperm antibodies (p less than 0.0001, versus fertile controls). Lymphocyte responses to phytohemagglutinin (PHA), concanavalin A (ConA) and pokeweed mitogen (PWM), however, were similar in 20 fertile and 136 infertile couples. Lymphocytes from infertile men with sperm antibodies had an enhanced stimulatory response to autologous sperm and seminal plasma. It is concluded that sperm antigens from autoimmune infertile men stimulate heightened immune responses to sperm antigens and altered distribution of white cell populations in both partners.     

Molecular aspects of mammalian fertilization

The initial communication between the gametes is a molecular,receptor-mediated process that takes place at the surface ofthe egg coat. The zona pellucida plays a central role in thisprocess such that, on the one hand, spermatozoa may bind toit and, on the other hand, it prevents polyspermy. In the mouse,ZP3, a glycoprotein of the zona pellucida with a mol. wt of84 kd, serves as a sperm receptor. Only a relatively small partof ZP3, namely certain O-linked carbohydrate side chains, isinvolved in the process of binding. These oligosaccharides probablybecome bound to enzymes associated with the plasma membraneof the sperm head and thus form an enzyme-substrate complex.A terminal -galactose has been found to be one of the decisivesugar molecules and, moreover, the critical chemical group.After sperm binding to the zona pellucida has taken place, thepolypeptide chain of ZP3 initiates the acrosome reaction inthe sperm head. In the mouse, numerous binding proteins havebeen detected in the sperm plasma membrane: these are enzymessuch as glycosyl transferases, proteinases, and glycosidases.A galactosyl transferase has been found on the surface of themouse sperm that binds specifically to N-acetylglucosamine inthe mouse zona pellucida. It is therefore apparent that carbohydrate-bindingproteins on the sperm surface mediate gamete recognition throughtheir high affinity and specificity for complex glycoconjugatesin the egg coat. In fact, it is not at all surprising that complementarycell-surface protein and glycoconjugates are involved in fertilization,since many somatic cells exhibit a similar mechanism of cellrecognition.     

Use of antisperm antibodies in differential display Western blotting to identify sperm proteins important in fertility

BACKGROUND: Antisperm antibodies (ASA) may be an important cause of infertility but current tests for the detection of ASA have poor prognostic value. The inadequacy of current tests may reflect the inability of these tests to define the antigenic specificity of the sperm proteins with which the ASA react. Identification of the sperm proteins that ASA bind to is a necessary preliminary step to the development of more useful diagnostic tests for ASA. METHODS: A sensitive Western blotting technique was used to compare the antigenic specificities of ASA from men who were infertile (n = 6) with those who were fertile following vasectomy reversal (n = 3). Normal fertile men (n = 3) and infertile men with known ASA (n = 4) were also included in the analysis. RESULTS: All men, including the normal fertile controls, had ASA detectable in our system. Several sperm proteins were identified that react with ASA from infertile but not fertile men. Quantitative differences in the binding of ASA to some proteins were also demonstrated. Additionally, we demonstrated that normal motile sperm are coated with an antibody that appears to be bound to sperm by a non-antigenic mechanism. CONCLUSION: Sera from all men contained ASA, but clearly some of these did not cause infertility. Characterization of the proteins that are antigens for ASA from infertile but not fertile men may allow the development of more accurate tests for infertility-inducing ASA. The significance of immunoglobulin G coated on normal sperm remains to be determined.     

Gene deletions in an infertile man with sperm fibrous sheath dysplasia

BACKGROUND: Asthenozoospermia may sometimes be related to geneticstructural defects of the sperm tail detectable by transmissionelectron microscopy. Dysplasia of the fibrous sheath (DFS) isa genetic sperm defect, characterized by dysplastic developmentof the axonemal and periaxonemal cytoskeleton. We report thecase of an infertile man with normal sperm count and total spermimmotility in which dysplasia of the fibrous sheath, Akap3,Akap4 gene deletions, meiotic segregation of chromosomes 18,X and Y and Y microdeletions were investigated. METHODS: A 32-year-oldman with a 3-year history of primary infertility presented atour Regional Referral Center for Male Infertility. Family medicalhistory, lymphocyte karyotype, PCR analysis, physical examination,hormone assays and semen analysis were performed. RESULTS: Ultrastructuralsperm evaluation showed dysplasia of the fibrous sheath. Immunostainingof AKAP4 protein was negative in sperm tails. PCR analysis revealedintragenic deletions of the Akap3 and Akap4 genes. Fluorescencein situ hybridization on sperm showed a high frequency of XYdisomy. CONCLUSION: In this infertile patient, our results suggesta possible relationship between dysplasia of the fibrous sheath,partial deletions in the Akap3 and Akap4 genes and absence ofAKAP4 protein in the fibrous sheath. These findings, however,were not detected in another four patients with dysplasia ofthe fibrous sheath. Our results require future confirmatorymolecular analyses.     

Application of proteomic methods for identification of sperm immunogenic antigens

Although the immunogenic properties of sperm have been explored for a few decades, none of the antigens studied so far appears to be an effective target, to inhibit the fertilization process or shown the full spectrum of sperm antigenic potential. Antisperm antibodies (ASA) collected from infertile individuals and prepubertal boys with cryptorchidism together with two-dimensional (2D) electrophoresis have been employed. Immunoreactive antigens were cored from silver stained 2D gels and analyzed by mass spectrometry (MS). The obtained sequences were searched in the published protein databases. Altogether, 35 different sperm entities were identified in accessible protein databases, out of which 10 appeared to be sperm-specific. Additionally, 6 amino acid sequences indicated novel (hypothetical) proteins. Seventeen sperm entities were detected in sera samples from immune infertile males and 18 entities in ASA-positive seminal plasma (SP). Interestingly, we identified a few sperm structures, none of them sperm specific in sera samples from infertile females. Although, infertile males from whom the ASA-positive SP samples were obtained, did not have ASA in their circulation, the range of sperm antigens detected by systematic and local antibodies overlapped to a great extent (six identical entities). Sera samples from prepubertal boys allowed to show antigens, previously thought to be only present on mature sperm. Three out of four detected were sperm-specific. Using serum and SP of ASA-positive infertile adults and sera samples of prepubertal boys with testicular failure, we have extended the range of known, immunogenic sperm proteins as well as identified some novel antigens (n=6) of human sperm for further characterization.     

Asthenozoospermia and the human sperm mid-piece

This study provides a quantitative comparison between surfaceand ultrastructural features of motile spermatozoa in asthenozoospermicand fertile men. The study group consisted of 10 individualswith persistent asthenozoospermia and the controls were 10 fertiledonors to a sperm bank. Scanning electron microscopy and imageanalysis were used to objectively measure sperm mid-piece andtail dimensions. Sperm mid-piece length was significantly shorter(P < 0.01) in asthenozoospermic subjects compared with thecontrols, with mid-piece width and tail length being comparable.Mid-piece ultrastructure was then examined with the transmissionelectron microscope and the number of mitochondrial gyres andtheir configuration recorded. At the ultrastructural level theasthenozoospermic subjects demonstrated significantly fewermitochondrial gyres (P < 0.001) than their fertile counterparts.Energy for sperm movement is provided by mitochondria and adeficit in these organelles in the sub-fertile cohort providesan explanation for poor sperm function in these subjects.     

A self-migration method for preparation of sperm for in-vitro fertilization

Human sperm samples (n = 211) were prepared for in-vitro fertilization(IVF) and embryo transfer by a self-migration procedure in Earle'smedium containing highly purified hyaluronic acid (Hya) (MW3 000 000) included to increase the viscosity of the medium.The method resulted in the recovery of a significantly higherpercentage of motile spermatozoa compared with the traditionalcentrifugation method, 87.5 ± 0.9% versus 76.1 ±1.3% (P < 0.001). When comparing media with and without Hyain the selfmigration method for preparation of normal spermsamples, the media containing Hya resulted in the recovery ofa significantly higher percentage of motile spermatozoa, 89.0± 0.8% versus 73.8 ± 2.0% (P < 0.001). In agroup of 80 consecutive couples entering our IVF programme,sperm samples from 44 of the men were allocated at random forthe self migration method in medium containing Hya and spermsamples from 36 men for preparation by centrifugation and swim-up.Significantly more pregnancies were achieved in the group preparedin medium containing Hya. It is concluded that self-migrationof sperm in a medium containing Hya is simple and rapid, andresults in a high recovery of motile spermatozoa which can beused for in-vitro insemination of human oocytes with favourableresults.