Test Yolk Research Articles

Flow cytometric analysis of enzymes in live spermatozoa before and after cryostorage.

Synthetic fluorogenic substrates, like the CellProbe reagents, can determine enzymes in vital human spermatozoa. These substrates will enter the cells without previous cell permeabilization and exhibit fluorescence after cleavage depending on enzyme activity. They consist of different peptide sequences, specific for the enzymes, and a fluorescein- or rhodamine 110-dye moiety. The number of positive cells and the intensity of the fluorescence can be determined by flow cytometric analysis. We investigated several enzymes (peptidases, proteinases, esterases, elastases and collagenases) in intact spermatozoa before and after cryoprotection. Semen samples with normal spermiogram parameters were cryoprotected using the freezing medium TEST yolk buffer (TYB). Fresh spermatozoa showed a marked fluorescence after incubation with the synthetic substrates for the aminopeptidase M, butyryl esterase, fluorescein diacetate (FDA)-and FDA/sodium fluoride (NAF)-esterase, ala-ala-pro-val (AAPV)-elastase, gly pro-leu-gly pro-(GPLGP)-collagenase, gly gly leu-(GGL)-subtilisin as well as lys-ala-(LA)-dipeptidyl peptidase (DPP) II. After cryopreservation the spermatozoal fluorescence increased applying substrates for butyryl esterase (P<0.05), prolyl-aminopeptidase (P<0.001) and val-lys-(VK)-cathepsin (P<0.001) most probably due to elevated enzyme activities. The activities of FDA-esterase (P<0.05) and FDA/NAF-esterase (P<0.05), AAPV-elastase (P<0.01), GPLGP-collagenase (P<0.05) and GGL-subtilisin (P<0.001) decreased after cryopreservation. The substrates for arg-gly glut-ser-(RGES)-elastase, gly phenyl-gly ala-(GFGA)-collagenase and threo-pro-(TP)-cathepsin were not cleaved before as well as after cryostorage. The substrates for subtilisin an

Freezing a few testicular spermatozoa retrieved by TESA

Men with azoospermia can now be treated using testicular sperm aspiration (TESA). New aspirations, in subsequent cycles, may be avoided using cryopreservation. Conventional sperm freezing techniques are not suitable for TESA samples with a small number of spermatozoa. Testicular spermatozoa were obtained from 10 azoospermic men undergoing TESA for a diagnostic objective. Two different freezing protocols were performed according to the number of spermatozoa found in the final suspension: between 100–2000, we used TEST yolk buffer with glycerol, adding it to testicular sperm (Method I); for less than 100, we injected them into cell-free human zona pellucida before adding a freezing medium (Method II). Sperm and motility recovery rates were 1% and 32.3%, and 88.2% and 26.6% for methods I and II respectively. The fertilisation rate was 13.3% and 23% for methods I and II respectively. This study represents our preliminary experience in freezing testicular spermatozoa collected by TESA. Preliminary observations show that it is possible to freeze a few testicular spermatozoa inside evacuated zona pellucida.

Birth of healthy male twins after intracytoplasmic sperm injection of frozen-thawed testicular spermatozoa from a patient with nonmosaic Klinefelter syndrome

Birth of healthy male twins after intracytoplasmic sperm injection of frozen-thawed testicular spermatozoa from a patient with nonmosaic Klinefelter syndrome

Use of a Novel HOS-PVP Reagent for Rapid Screening of Immotile Human Sperm for Membrane Integrity

Objective: For ICSI, motile spermatozoa represent the safest choice for successful egg activation and normal embryo development. However, in some cases, only immotile sperm are available, which makes viable sperm selection problematic. The objective of this work was to develop a rapid and simple technique to easily screen practical numbers of immotile spermatozoa individually using micromanipulation by incorporating polyvinylpyrolidine into a hypoosmotic buffer (HOS-PVP). Design: Prospective. Setting: Andrology and IVF Laboratory. Patients: Men (n=39) presenting for semen analysis for infertility evaluation. Mean Age: 37.7 (range: 28∼50); Mean sperm concentration: 163.2 (range 12∼600); Mean motility: 84.3 (range: 26∼89); Mean morphology: 35.0 (range 15∼53). Main outcome measure(s): Using an ICSI pipette, micromanipulated individual human spermatozoa (1–10) were transferred from 10% PVP in isotonic buffer into an adjacent drop of 10% PVP in HOS buffer. Using 400× magnification spermatozoa were rapidly scored for an immediate response (1–10 seconds) as swelling or “curling” of the terminal 5–10 microns of the tail tip. Individual motile and immotile spermatozoa from 39 specimens were compared before and after cryopreservation in conventional TEST yolk buffer. Results: A total of 1000 individual spermatozoa were analyzed from 39 individuals. As expected from work of previous investigators using HOS reagent alone, the vast majority of motile spermatozoa show immediate tail tip swelling upon exposure to HOS-PVP (mean=89.1%, range=52%∼100%). However, the novel inclusion of PVP into the HOS solution enables a practical evaluation of many sperm (up to 20) simultaneously since they remain stable in the visual fields. Once identified, individual spermatozoa can be rapidly returned within seconds into standard isosmotic PVP for subsequent use in ICSI. There were significant differences in tail tip curling between both fresh and thawed immotile sperm in HOS-PVP. While only 8.96% (SD=1.88; range=0∼20%) of individual fresh immotile sperm showed tail tip curling, significantly more 24% (SD=2.83; range 0∼60%) of those screened after thawing showed this curling (P=0.004916; Chi Square). This difference is probably due to the subpopulation of motile sperm whose motility did not survive the thaw but whose membranes were still intact. Identification of this population is useful when choices between only a few immotile spermatozoa must be made, either fresh or after thawing. Preliminary observations suggest that immotile sperm selected in this way activate mouse oocytes after injection using ICSI than randomly selected immotile spermatozoa. Conclusion: A HOS-PVP reagent can be used to rapidly screen for and to choose individual immotile sperm for use in ICSI.

Comparison of protective media and freezing techniques for cryopreservation of human semen

Objective: To evaluate the influence of cryopreservation medium and freezing–thawing techniques on human sperm motility and morphology. Study design: 63 semen samples were obtained from 39 donors to the artificial insemination programme. Possible effects of the sperm dilution with cryomedium on the motility were examined 10 min after exposure of 24 high initial quality semen samples to TEST–yolk {zwitterion–citrate–egg yolk extender containing TES [N-Tris (hydroxymethyl) methylaminoethane sulfonic acid] and Tris [(hydroxymethyl) aminomethane]} and human sperm preservation medium (HSPM). Post-thaw sperm motility from 24 frozen semen samples was examined comparing the cryoprotective efficacy of TEST–yolk and HSPM following different freezing techniques (vapour freezing, fast programmable freezing and slow programmable freezing). The relationship of sperm morphology to the effects of freezing was investigated on 39 semen samples following different freezing techniques. Post-thaw sperm motility from 39 frozen semen samples was compared among three groups divided according to the percentage of morphologically normal cells (<40, 40–50 and >50%) in fresh semen. Results: Exposure of spermatozoa to cryomedia for 10 min at room temperature significantly reduced motility in TEST–yolk treatment group for 9% and in HSPM treatment group for 18% (P<0.01). The recovery of motile sperms (mean±standard deviation) was 49±15.7, 43±15.2 and 52±16.8% when TEST–yolk was used and 34±17.8, 32±18.2 and 50±13.6% when HSPM was used as a cryopreservative following vapour freezing, and fast and slow programmable freezing, respectively. Following vapour freezing and also following fast programmable freezing, the recovery of motile sperm was significantly higher (P<0.05) after addition of TEST–yolk medium than after addition of HSPM. Post-thaw motility of the sperm cryopreserved in HSPM showed significant differences (P<0.05) after three different freezing techniques. The recovery of motile sperms was 57±26.4, 38±8.6 and 38±17.3% in groups with >50, 40–50 and <40% morphologically normal cells, respectively. The percentage of morphologically normal spermatozoa was reduced 8% after vapour freezing and 6 and 3% after fast and slow programmable freezing, respectively. The results were statistically analysed using sas/stat software. Conclusions: Slow programmable freezing was superior to vapour freezing and fast programmable freezing as a method for sperm cryopreservation. However, none of these methods of freezing had discernible effects on sperm morphology. Motility of spermatozoa decreased due to the exposure of semen to cryomedium. TEST–yolk was a superior cryomedium to HSPM. Fresh semen with more than 50% of morphologically normal cells showed the best recovery of motile cells after freezing and thawing.

Hidden effects of cryopreservation on quality of human spermatozoa.

The effects of cryopreservation on two characteristics of human spermatozoa were investigated: the early phases of disturbed plasma membrane function and the activity of enzymes in intact spermatozoa. The membrane function was detected by means of the calcium-dependent binding of fluorescein isothiocyanate (FITC)-conjugated Annexin V to sperm plasma membranes. Annexin V monitors the translocation of phosphatidylserine from the inner to the outer leaflet of the plasma membrane, which is one of the earliest features of membrane disintegration. For the second aim synthetic fluorogenic substrates for peptidases, proteinases, esterases, elastases and collagenases were applied. These substrates, CellProbe trade mark reagents consist of different peptide sequences, specific for the enzymes, and a fluorescein- or rhodamine 110-dye moiety. They enter the cells without previous membrane permeabilisation and exhibit fluorescence after cleavage depending on enzyme activity. The number of positive cells and the intensity of the fluorescence were determined by flow cytometric analysis comparing fresh spermatozoa with cryopreserved ones. Thirty-five semen samples collected from 35 donors were cryopreserved using the freezing medium TEST yolk buffer. All specimens showed normal spermiogram parameters. Twenty-five of the samples were used for detection of Annexin V-FITC binding and 10 semen samples for investigations of the intracellular enzymes. The Annexin V-assay applied two fluorescent dyes (Annexin V, AN and propidium iodide, PI) which led to three groups of spermatozoa (a) viable spermatozoa (AN V-negative and PI-negative), (b) dead spermatozoa (AN V-positive and PI-positive) and (c) cells with impaired but integer plasma membrane (AN V-positive and PI-negative). The percentage of vital Annexin V-negative spermatozoa (x +/- SEM) decreased significantly (p < 0.001) from fresh spermatozoa (51.6 +/- 3.1) to cryopreserved spermatozoa (26.6 +/- 2.2%) and was associated with their motility (57.9 +/- 1.9% motile fresh spermatozoa vs. 22.6 +/- 3.9% motile sperm after cryopreservation). Of the spermatozoa 28.2% were Annexin V-positive before and 44.4% after cryostorage even though they did not bind to PI. Thus, vital spermatozoa showed a disturbed membrane function indicating viability before as well as after cryostorage. Moreover, after cryopreservation the spermatozoal fluorescence increased applying substrates for butyryl esterase (p < 0.05), prolyl-aminopeptidase (p < 0.001) and val-lys-(VK)-cathepsin (p < 0.001). In contrast, the activities of fluorescein diacetate (FDA)- and FDA/sodium fluoride (NAF)-esterase (p < 0.05), ala-ala-pro-val-(AAPV)-elastase (p < 0.001), gly-pro-leu-gly-pro-(GPLGP)-collagenase (p < 0.05) and gly-gly-leu-(GGL)-subtilisin (p < 0.001) decreased after cryopreservation. The substrates for arg-gly-glut-ser-(RGES)-elastase, gly-phenyl-gly-ala-(GFGA)-collagenase and threo-pro-(TP)-cathepsin were not cleaved before as well as after cryostorage. In addition to the known effects of sperm cryopreservation our results showed two further alterations of human ejaculated spermatozoa: (a) disturbed plasma membrane function, which is not detectable by supravital staining and (b) a changed pattern of intracellular enzyme activities.

Effect of storage temperature on sperm cryopreservation

Objective: To evaluate the influence of cryopreservation temperature on human sperm motility and morphology. Design: Controlled study, investigator was blinded to the type of cryopreservation. Setting: University-based andrology laboratory. Patient(s): Sixteen semen samples with normal motility and sperm count from men after a fertility work up. Intervention(s): Semen aliquots were either stored in a mechanical freezer at −70°C or in liquid nitrogen at −196°C for 7 days or 3 months. Test yolk buffer was used as a cryoprotectant. With use of a programmable freezing unit, all samples were cooled at a controlled rate. Main Outcome Measure(s): Sperm motility and morphology. Result(s): After 7 days of cryopreservation, there was a greater decrease in sperm motility among specimens maintained at −70°C than among those maintained at −196°C (47% versus 39% decrease). The difference in sperm motility was even greater after 3 months of cryopreservation (72% versus 39% decrease). No difference in postthaw sperm morphology was detected among sperm preserved at −70°C versus −196°C. Conclusion(s): Sperm cryopreservation at −196°C is superior to cryopreservation at −70°C. Sperm can be stored at −70°C for a short period of time with a relatively modest loss of motility.

Reactive oxygen species generation by seminal cells during cryopreservation

Objectives. To test the hypothesis that conventionally used procedures for semen cryopreservation may cause an increase in the production of reactive oxygen species (ROS) by sperm or by seminal leukocytes, which may contribute to poor sperm function following cryopreservation. Methods. Eighteen semen specimens with normal parameters from healthy male donors 22 to 40 years of age were each divided into two portions. The first portion was combined 1:1 with Test Yolk Buffer-Glycerol Freezing Medium and was frozen by gradual cooling into liquid nitrogen (−196°C). The second portion was washed and the cells were resuspended in Sperm Washing Medium (SWM) and incubated at room temperature to serve as controls. After a period of treatment, frozen samples were thawed and semen cells were washed and resuspended in SWM. ROS generation by semen cells from each treatment group was measured on a luminometer. Sperm motility, sperm viability, and sperm membrane integrity were also measured in both control and freeze-thaw samples. To further assess ROS generation by semen cells during the cooling process, aliquots of washed semen cells and purified polymorphonuclear leukocytes (PMNs) were incubated separately at different temperature conditions (37°C, 22°C, 4°C, and −20°C). ROS activity in each treatment group was measured and compared with each other. Results. In both semen cells and PMNs, ROS activity increased significantly during the cooling process. The highest ROS levels were recorded in both groups when cooled to 4°C. The ROS levels were extremely low in samples cooled to −20°C and in freeze-thaw samples, probably due to marked loss of cell viability. Conclusions. Gradual reduction of temperature during the process of semen cryopreservation can cause a significant ROS generation by semen cells. ROS is particularly elevated during cooling if the semen sample is contaminated by more than 0.5 × 10 6 leukocytes. Removal of leukocytes from semen samples or treatment with antioxidants prior to cryopreservation may improve sperm viability and function.

Correlation between intact sperm acrosome assessed using the Spermac stain and sperm fertilizing capacity.

Accurate determination of sperm acrosomal status is important in fertility studies. The objective was to correlate the percentage of intact acrosome assessed using the new Spermac stain with the capacity of sperm to fertilize oocytes. Sperm specimens were processed either by centrifuge wash, 48:95 Percoll gradient or test yolk buffer (TYB) wash, and tested using the zona-free hamster oocyte assay. The results indicated a correlation between the percentage of sperm with intact acrosome reaction and the percentage of sperm penetrating the oocytes in the TYB-washed group. The data suggest the usefulness of the Spermac stain for assessing the acrosomal status and in predicting the fertilizing capacity of the sperm.

Effect of cryopreservation on semen quality in patients with testicular cancer

Objectives Current techniques in cryopreservation of human semen substantially decrease sperm quality. In addition, the pregnancy rate using cryopreserved sperm obtained from testicular cancer patients is lower than when sperm from normal fertile men is used. However, it is still unclear whether cryopreserved sperm from these patients is inherently defective or if the sperm loses its motility after thawing. This study was undertaken to assess the effect of cryopreservation on the quality and motion characteristics of semen from patients with testicular cancer before definitive therapy compared with semen from normal volunteers. Methods We compared the sperm quality before and after cryopreservation in samples from 34 patients with testicular cancer and 30 normal volunteers who were referred for sperm banking over a 7-year period. The effects of cancer stage and histologic type on various semen parameters were also examined. A computer-assisted semen analysis was performed before and after cryopreservation on each specimen. The nitrogen vapor technique using Test yolk buffer with glycerol as a cryoprotective agent was used for cryopreservation. The motile sperm count and motion characteristics (motility, velocity, linearity, amplitude of the lateral head movement, motility index) were analyzed before and after cryopreservation and compared between the groups. Results Semen quality did not significantly differ among patients with Stage I, II, or III cancer. However, semen quality tended to be poorer at higher cancer stages. In general, semen quality was better among patients with pure seminomas than with pure embryonal tumors; quality was worst among patients with mixed germ cell tumors. However, 71.4% of patients with mixed tumors presented with Stage III disease, whereas all patients with seminomas presented with Stage I disease. Significant differences were also seen in prefreeze motility (median, 42% [interquartile range, 24 to 51] versus 60.5% [range, 49 to 73]; P = 0.0004) and motile sperm count (6.7 × 10 6/mL [range, 3.4 to 14.4] versus 50.0 [range, 24.6 to 72.0]; P = 0.0001) in patients compared with controls, respectively. The motile sperm count and percent motility significantly decreased in both patients and controls after cryopreservation ( P = 0.0001). However, the percentage decline in motile sperm count and motion characteristics after cryopreservation did not differ significantly between patients and controls ( P > 0.01). Conclusions We conclude that the effect of cryopreservation on sperm quality in patients with testicular cancer is identical to its effect on sperm from normal fertile men. Differences in values after preservation are explained by poor semen characteristics before freezing; semen quality declines with more extensive disease. Stage I patients also had poorer quality than control subjects. Thus, we recommend that routine sperm banking be encouraged among all patients with testicular cancer before the initiation of specific medical treatment. We also recommend that future efforts be focused on improving the technique of sperm banking.

Effect of caffeine and pentoxifylline on human sperm motility after one week storage at 3 degrees C

The effect of caffeine (C) and pentoxifylline (P) on the sperm motility was examined after a 3 degrees C storage in a TEST yolk buffer for one week. Thus stored 24 semen samples from infertile patients were incubated with or without C or P at 37 degrees C. The motility was analysed for up to 240 minutes. The poststorage addition of C (1 mM) and P (1 mM) significantly increased the serum motility rate (p < 0.01). The stimulant effect of C and P was rather transient lasting no more than 2 hours. The prestorage addition of C and P in a preservant TEST yolk buffer was less effective than the poststorage administration. According to the time-related profiles of the sperm motility, 24 cases were divided into three groups. In group A (11 cases), the sperm motility rate increased from 30 to 90 minutes even without C or P. This group was responder of C and P. In group B (8 cases), the motility rate increased only in response to C and P and, in group C (5 cases), no such responses were observed. The prestorage semen qualities in terms of the sperm concentration, the motility rate, the velocity, the linearity, and the amplitude of the lateral head displacement were compared in these three groups. There was a significant difference between these 3 groups (p < 0.05) on the parameters in terms of the sperm concentration, motility rate and velocity. It was suggested that these prestorage parameters predict the sperm activity in response to C and P after a one week 3 degrees C storage.

Failure of test yolk buffer to decrease antisperm antibodies on sperm.

The objective of this study was to evaluate whether the suspension of sperm naturally bound or artificially coated with antisperm antibody (ASA) in test yolk buffer (TYB) will reduce the percentage of sperm positive for ASA. Attempts were made to repeat a study performed by Lam et al., in which they claimed a significant decrease in percentage of sperm positive for both IgA and IgG. Furthermore, the study attempted to determine if this methodology would similarly reduce the percentage of sperm naturally bound with ASA. There was no reduction in postincubation levels in percentage of sperm having attachment of IgA or IgG in either the sperm specimens naturally bound with ASA or donor sperm artificially coated with antibodies. Incubation in TYB does not appear to be an effective method to reduce the percentage of sperm bound with ASA. Possibly, it could reduce the amount of ASA bound to each sperm, but this would have to be proven by other studies.

Synergistic effect of TEST--yolk buffer treatment and glass wool filtration of spermatozoa on the outcome of the hamster oocyte penetration assay.

Enhanced penetration of zona-free hamster oocytes occurs after spermatozoa have either been treated with TEST--yolk buffer or processed by glass wool filtration. The present study was conducted to determine if a synergistic effect on fertilizing ability could be achieved by combining these two treatments. Ejaculates (n = 16) were treated with TEST--yolk buffer (TYB) and divided into two aliquots. One aliquot was kept as control and the other aliquot was filtered through glass wool prior to the sperm penetration assay. The TYB--glass wool filtered spermatozoa penetrated a significantly higher percentage of oocytes than the TYB treated spermatozoa.

Composition, Functionality, and Sensory Evaluation of Eggs from Hens Fed Dietary Menhaden Oil

ABSTRACTEnrichment of omega‐3 fatty acid content of egg yolk may increase consumer acceptance of egg products if eggs maintain characteristic functionality, exhibit compositional stability, and are sensorially acceptable. The diet of laying hens was enriched with 3% menhaden oil. Arachidonic acid (20:4n‐6) was decreased 70.2%, and linolenic (18:3n‐3) and docosahexaenoic acid (22:6n‐3) were increased 78.5% and 356%, respectively, in egg yolk. Eicosapentaenoic acid (20:5n‐3) was also incorporated into test egg yolk as compared to nondetectable levels in control eggs. Cooking did not alter the fatty acid composition of eggs nor were functional properties of test eggs affected. Panelists differentiated n‐3 enriched eggs from controls (P ≤ 0.01) when scrambled but not when hard cooked.

Comparative study on the use of human follicular fluid or egg yolk medium to enhance the performance of human sperm in the zona-free hamster oocyte penetration assay

A prospective study was conducted on 61 unselected semen samples from infertile patients to compare the efficacy of human follicular fluid (hFF) or TEST-egg yolk medium treatment in enhancing sperm performance in the zona-free hamster oocyte penetration (HOP) test. Both hFF and TEST-egg yolk enhanced penetration scores significantly in the HOP test in comparison with the control swim-up sperm preparation. The TEST-egg yolk medium treatment, however, exhibited the highest penetration scores, and there were higher percentages of preparations passing the 10 or 20% penetration rate cut-off point with sperm exposed to the TEST-egg yolk medium than those exposed to hFF. These results indicate that modification of the original HOP-test protocol by incorporation of TEST-egg yolk medium gives a greatly increased penetration rate when compared to the addition of hFF in such a system.

Penetration of zona-free hamster oocytes by ejaculated cryopreserved gorilla spermatozoa

Semen obtained by electroejaculation from two lowland gorillas were cryopreserved in TEST yolk to evaluate the ability of spermatozoa to penetrate zona-free hamster oocytes. Thawed semen was processed through a two-layer Percoll density gradient to obtain motile spermatozoa for the SPA. The processed sperm penetrated greater than 25% of the zona-free hamster oocytes. Thus, the use of TEST yolk to cryopreserve gorilla semen and processing the thawed semen through Percoll gradient to concentrate motile spermatozoa may facilitate sperm capacitation and the ability to penetrate oocytes.

Sources of variation in the computer-assisted motion analysis of rat epididymal sperm

Random and nonrandom factors associated with sample preparation and the automated analysis (CellSoft) of rat cauda epididymal sperm motion were studied. Random factors included inherent system variation at both the individual cell level and at the multiple cell level. Repeated analyses of identical tracks across grey level revealed a statistical interaction between grey settings and curvilinear velocity. However, in multiple track analyses, grey level was seen to be a factor only at higher settings. Nonrandom factors included time after sample preparation, dilution medium, and sample preparation procedures. Using a nicked preparation of the entire cauda epididymis from Long-Evans rats, the effects of time were studied on sperm suspended in 1) phosphate-buffered saline + 10 mg BSA/mL, 2) TEST yolk buffer, and 3) Medium 199. In PBS/BSA, the percent motile sperm estimate decreased (50% to 30%) over an hour, while the curvilinear velocity increased (127 to 142 μm/sec). Both sperm motion parameters were maintained in the TEST yolk buffer and in the Medium 199, although at lower values for the latter. Evaluation of the relative contribution of several factors, nested within sample, to the overall variance of three separate motion endpoints revealed that there was a large variation from field to field, negligible variation between overall CellSoft analyses of 200 cells or more, low variation at the preparation aliquot level, and moderate variation at the animal level. In planning experiments to test for effects on sperm motion endpoints, consideration of the relative contribution of the individual study factors to the overall variance of the parameter estimates will result in more sensitive experimental designs.

Treatment of human spermatozoa with an egg yolk medium can enhance the outcome of in vitro fertilization

In at least 4 of 7 cases, fertilization of intact human oocytes was more successful when spermatozoa were pretreated with TEST yolk medium at 5 degrees C for 2 hours as compared with the standard treatment with Ham's F-10 only. Both pregnancies that were obtained after the transfer of the fertilized oocytes resulted from oocytes fertilized by TEST yolk-treated spermatozoa. No decrease in fertilization occurred in any of the cases after TEST yolk treatment. If these results hold true for a larger series of patients, it may be worthwhile for the standard IVF incubation system of spermatozoa to include TEST yolk.

TEST-Egg Yolk Buffer Storage Increases the Capacity of Human Sperm to Penetrate Hamster Eggs in Vitro

Departments of Obstetrics and Gynaecology and Medicine, University of Hong Kong, Queen Mary Hospital, Hong Kong Int. J. Androl. 40: 517, 1987

TEST-Egg Yolk Buffer Storage Increases the Capacity of Human Sperm to Penetrate Hamster Eggs in Vitro

TEST-Egg Yolk Buffer Storage Increases the Capacity of Human Sperm to Penetrate Hamster Eggs in Vitro