F. Le R. Fourie

Multiple attempts at embryo transfer: effect on pregnancy outcome in an in vitro fertilization and embryo transfer program.

PurposeThis study derives from the observation that a correlation exists between failed first attempts (FFA) at embryo transfer caused by one or more embryos remaining in the catheter and reduced pregnancy rates (20.3 vs 3.0%). The aim of this study was to examine the relationship between failed first attempts at transfer and contamination of the transfer set; the related aspects of cervix dilatation and late embryo transfer were also investigated.ResultsThe following observations were made. Retention of embryos in the transfer sets significantly reduced the pregnancy rate (P =0.015); catheters contaminated with blood and cervical mucus indirectly contributed to this effect by increasing the incidence of failed first transfer attempts. Even though cervical dilatations, if indicated by uterus sounding, were done 2 days before embryo transfer, no pregnancies were effected in these 18 cases (P =0.0001). Late transfers of embryos, due to delayed fertilization or slow cleavage rates, yielded a pregnancy rate of 10.5%.ConclusionThe approach of immediately retransferring retained embryos does not solve the problem of reduced pregnancy rates in FFA cases. It is suggested that ET should be repeated 1 day later in FFA cases in an attempt to improve pregnancy rates.

Microbial flora in semen during in vitro fertilization

Semen samples from 183 consecutive unselected men participating in an in vitro fertilization program were retrospectively studied to determine the bacterial and fungal contamination rate before and after antibiotic treatment. To ascertain the influence of semen preparation (wash and swim-up method) on the incidence of microorganisms, semen from 102 male patients was studied before and after swimup. Antimicrobial treatment by, prescription of antibiotics decreased the incidence of pathogens by 16.3% (P<0.0001). Semen processing was more effective by ridding 57.4% of semen samples of microbial contaminants (P<0.0001). When infection of culture media was observed during routine microscopy, all infected oocytes were degenerated, without evidence of fertilization or pronuclei.

Primate in vitro fertilization research: preliminary results on the folliculogenic effects of three different ovulatory induction agents on the chacma baboon, Papio ursinus

1. A study was conducted on the chacma baboon, Papio ursinus with three ovulation induction agents in an effort to define a preferential stimulatory protocol with regards to the number and quality of oocytes obtained. 2. Three folliculogenic agents applied in four stimulatory protocol regimens comprised clomiphene citrate in a high (100 mg/day) and low (50 mg/day) dosage, a combination of clomiphene citrate and pregnant mare serum, and human menopausal gonadotropin. 3. A total of 159 oocytes were aspirated by laparotomy from 10 baboon females in 20 induced cycles with an average of 8.0 +/- 5.4 oocytes per aspiration. 4. The highest mean number of oocytes (11.3 +/- 6.7) were obtained with the clomiphene/pregnant mare serum gonadotropin combination. 5. The best fertilization rate was obtained with clomiphene 50 mg. 6. The highest incidence of oocytic cleavage and embryo transfer were achieved with human menopausal gonadotropin (14.8%).

Comparative haematology of some South African birds

Abstract Haematological and biochemical parameters of 251 birds representative of 26 species and six orders of the Class Aves were investigated.

Spectrophotometric absorbance of follicular fluid: A selection criterion

PurposeThe aim of this study was to ascertain the absorbance profiles of uncontaminated follicular fluids (FF) vs blood and medium contaminated FF, and to test the hypothesis that blood contamination alters the biochemical status of FF.MethodsFF (n =655) from 230 patients were scanned spectrophotometrically during an ongoing in vitro fertilization (IVF) program. Based on spectrophotometric evaluation, blood contaminated and uncontaminated FF (n =39 patients/FF) were analyzed for electrolyte content, inorganic phosphate, glucose, creatine, urate, total protein, albumin, bilirubin, cholesterol, alkaline phosphatase, γ-glutamyltransferase, alanine aminotransferase, aspartate transminase, and lactate dehydrogenase by means of a SMAC analyzer.ResultsLaparoscopic aspiration resulted in the collection of significantly more clear FF compared to the transvaginal procedure (P =0.001). Uncontaminated fluid depicted a single mean absorbance at 458.0 nm. Blood contaminated FF could be identified by spectrophotometry and revealed three peaks at 418.12, 540.13, and 575.32 nm, respectively, according to the degree of blood contamination. Follicular fluid diluted with Earles Balanced Salt Solution (EBSS) displayed an additional mean peak at 561.4 nm. Potassium, glucose, and all of the above mentioned enzymes revealed significantly higher levels in blood contaminated FF (P < 0.05).ConclusionsFrom this study, it is concluded that blood contamination and dilution with culture medium influence the biochemical composition as well as the absorbance spectrum of follicular fluids. This procedure is advocated as a prerequisite before quantifying FF content.

Supplementation of Ham's F10 culture medium with three different sera in the culturing of baboon oocytes

1. Ten female baboons (Papio ursinus) were stimulated for a total of 20 cycles with 3 ovulation induction agents. 2. Oocytes obtained were randomly allocated to Hams F10 culture medium supplemented with human fetal cord serum, primate serum or commercial fetal bovine serum respectively. 3. Fertilization occurred (38.1-45.5%) in all 3 supplements, but cleavage and embryo development was more successful in culture medium supplemented with fetal bovine serum. 4. Eight embryos were cultured and 6 (75%) of these were cultured in fetal bovine serum supplemented medium.

Spectrophotometric analysis of human follicular fluid with regard to in vitro fertilization (IVF) parameters, follicular protein, and hormone content

PurposeOur purpose was to investigate possible relationships with spectrophotometric absorbance (458-nm region) and biochemical variables in follicular fluid (FF) as well as in vitro fertilization (IVF) outcome.MethodsThis study included 227 normal ovulatory women undergoing oocyte retrieval for IVF. Blooduncontaminated fluid samples, identified by spectrophotometry, were investigated. Spectrophotometric absorbance of FF at 458 nm (n = 426), as well as hLH, FSH, PRL, hCG, testosterone, sialic acid, α1-antitrypsin and plasminogen of selected fluids, was analyzed.ResultsSmall-volume follicles (≤2 ml) were associated with higher absorbance profiles (P <0.05), when compared to volumes greater than 2 ml. Our data suggest that the presence or absence of an oocyte, the potential of an oocyte to fertilize or cleave, failed to show any relationship with maximum FF absorbance at 458 nm. Maximum absorbances were significantly lower in FF from patients who subsequently became clinically pregnant (P =0.039). No correlation between FF absorbances and biochemical parameters (P >0.15) were established.ConclusionsAbsorbance of clear FF at 458 nm should not be viewed as the single parameter to predict oocyte development in vitro.

A preliminary study of the fine structure of in vitro fertilized human embryos

The fine structure of rejected human in vitro fertilized embryos was studied by light and transmission electron microscopy. Apart from normal cytoplasmic features, changes suggestive of degeneration are discussed. Cytoplasmic blebs and cellular debris as well as primitive interblastomeric junctions are also described.

Electron microscopic study of human sperm membrane isolation

ObjectOur purpose was to isolated pure, homogeneous human sperm membranes, free of cellular contaminants.MethodsDonor semen samples collected after masturbation were stored at −70°C and eventually pooled. Each attempt at sperm membrane isolation required 800 × 106 spermatozoa which were sonicated by ultrasound (40% output; Vibra Cell). The effect of sonication time (3 × 5, 3 × 15, and 180 sec) on membrane isolation was investigated. Sonicated samples were centrifuged (500g, 5 min) and the supernatant was pipetted off. The supernatant of the centrifuged sample was layered on either a sucrose cushion (supernatant on 1.6 M sucrose) or a discontinuous sucrose gradient and centrifuged (100,000g, 1 hr). Contents of supernatants of sonicated samples and fractions (sucrose interfaces) were then fixed in 1.0% tannic acid and 2.5% buffered glutaraldehyde and examined electron microscopically using standard procedures.Results(1) The optimal sonification time was found to be 3 × 15 sec. (2) Membrane isolation using a sucrose cushion was found to be inadequate, showing significant cellular contamination. (3) Sperm membrane isolation from the sucrose interface between 0.75 and 1.05 M sucrose was found to be most effective.ConclusionThe advantage of this method is its simplicity. The drawback of this method is the large number of spermatozoa required for membrane purification.

Successful transcervical embryo transfer in two strains of mice

Twee muisrasse (’n swart Balb/C x C5 BL-ras en ’n wit NMRI-ras) is tot superovulasie gestimuleer en daarna toegelaat om met mannetjies van dieselfde ras te kopuleer. Die swart skenkermuise is 3,5 dae na kopulering deur servikale dislokering gedood en ’n totaal van 460 blastosiste is vanuit die fallopiaanse buise gespoel. Ses blastosiste afkomstig van die swart skenkerras is transservikaal in elk van die 71 wit ontvangermuise teruggeplaas 2,5 dae na kopulering. ’n Totaal van 51 geboortes na terugplasing is gedokumenteer met 31,4% voorkoms van swart fenotipiese afstammelinge.