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Biomedical subjects

T F Kruger

Publications and source records attributed to T F Kruger.

At least 127 records · Page 7Linked to original sources
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In vitro fertilization failure: identification of gamete defects by investigation of sperm-zona pellucida binding capacity of unfertilized oocytes.

The objective of the study was to determine whether fertilization failure was due to spermatozoal or oocyte factors. Twenty-five unfertilized oocytes from 12 IVF/GIFT couples showing total or partial fertilization failure were evaluated for sperm zona binding potential under hemizonaeassay (HZA) conditions. Hemizonae were separately incubated with a sperm sample from the husband and that of a fertile control. Tight sperm binding to hemizonae was assessed. First, among the 12 patients, results showed a possible zona defect thought to be the cause of fertilization failure in five cases. Second, in two cases, fertilization failure was possibly caused by poor sperm binding potential of spermatozoa. Third, in two cases, fertilization failure was possibly caused by an oocyte defect, and fourth, three cases showed a mixture of possible causes. The results stress the need to develop a sequential analytic programme for those couples with repeated total or partial fertilization failure.

Adult↗
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Comparison of two staining and evaluation methods used for computerized human sperm morphology evaluations.

The purpose of the study was to analyse the agreement between computer analysed (Hamilton Thorne, IVOS Dimensions Version 3) normal sperm morphology and values obtained from 97 slides stained according to the Papanicolaou and Diff-Quik method. Liquefied semen samples were washed once by centrifugation and air dried smears on slides were made, which were stained according to the Papanicolaou and Diff-Quik method and analysed by computer. The paired t-test was used to assess whether any bias existed between the two methods. The limits of agreement were calculated using the Bland and Altman approach and a modification of this approach (mean-dependent limits). A significant bias of 1.6% was obtained in favour of higher normal sperm morphology percentages when using the Diff-Quik method. The standard limits of agreement were -13.4% to 16.6%, whereas the mean-dependent limits of agreement were 1.6% [5.8 + 0.6 (mean percentage normal morphology)]. Statistically, the Diff-Quik and Papanicolaou staining methods produce different normal sperm morphology profiles. These inherent differences may, therefore, require the establishment of new normal sperm morphology thresholds for male fertility, based on clinical data, when using the Diff-Quik staining method in conjunction with computerized analysis.

Azure Stains↗
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Effects of different staining and washing procedures on the results of human sperm morphology evaluation by manual and computerised methods.

The purpose of this study was to investigate the effect of different staining and washing procedures on the results of human sperm morphology evaluation by manual and computerised methods. Furthermore, it was intended to find the staining and washing combination which would provide optimal readability for computer-assisted sperm morphology evaluations. In phase one, four staining methods were evaluated for smears prepared from the resulting samples following a two times washing procedure. In phase two, 20 semen samples were used to compare the Diff-Quik and Papanicolaou staining methods, following one and two washes. All manual readings, of Papanicolaou and Diff-Quik stained smears, were comparable with each other, with means between 7.3% and 7.9% normal spermatozoa. All the manual readings were also comparable to the computer readings of the Diff-Quik slides following one and two washes with means of 9.0% and 5.9%, respectively. However, due to the higher computer readings found for the Papanicolaou stained smears, with means of 13.9% and 13.5% following one and two washes, respectively, a statistically significantly difference between overall computer and manual readings was found (Wilks' Lamda, P = 0.0002). Taking all data into consideration, it could be concluded that the one wash Diff-Quik stained smears was the optimal preparation method for computerised sperm morphology evaluation, comparing favourably with manual evaluations.

Evaluation Studies as Topic↗
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Effect of ethanol on semen characteristics of vervet monkeys.

The effect of alcohol on spermatogenesis and sperm morphology has been of interest to many andrologists. Six adult male vervet monkeys were utilized to evaluate the effect of ethanol on semen quality. A decrease of the percentage of normal spermatozoa was observed after 10 months of exposure to alcohol.

Animals↗
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Effect of lead and organophosphates on sperm morphology.

Lead and organophosphates can exert a toxic action on the male reproductive function. A cross-sectional study involving 40 men was undertaken to investigate the effect of these toxins on sperm morphology. Based on the percentage of morphologically normal spermatozoa, subjects were divided into two groups. The control group (sperm morphology greater than or equal to 20% normal forms) consisted of 18 patients, and the study group (sperm morphology less than 5% normal forms) consisted of 22 patients. Lead, pseudocholinesterase, dibucaine, and fluoride levels were measured in blood samples of these men. The 22 men with low morphologically normal forms (less than 5%) did not display significantly different levels of the abovementioned substances compared to the control group.

Butyrylcholinesterase↗
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Ultrastructural parameters of fertile cryopreserved human sperm.

Two hundred and forty-four cryopreserved semen samples were used for artificial insemination by donor (AID). All samples were examined for motility and concentration. Thirty of the samples resulted in pregnancies. These samples were further examined ultrastructurally. There was no difference in sperm motility or concentration between the samples that did or did not result in a pregnancy. The ultrastructural characteristics of the samples that resulted in pregnancies revealed that only 15% of sperm (SD = 7.67) possessed normal morphology and had undamaged acrosomes after cryopreservation.

Acrosome↗
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Effect of semen characteristics on pregnancy rate in a gamete intrafallopian transfer program.

The aim of this study was to evaluate the influence of sperm morphology, swim-up concentration, and insemination volume on pregnancy outcome in patients undergoing gamete intrafallopian transfer (GIFT) treatment in whom the male partner had a morphology of less than 14%. Only patients who received four oocytes were entered into this study (n = 103). In all cases the swim-up procedure time was standardized to 1 h and the insemination concentration was standardized to 500,000 per oviduct. There was no significant difference in pregnancy rate when normal morphology, swim-up concentration, or insemination volume were used as predictors. In the P pattern group (< 4% normal forms) only four of the 28 (14%) patients had ongoing pregnancies, whereas in the G pattern group (5% to 14% normal forms) 16 out of 75 (21%) had ongoing pregnancies (P vs G, p > 0.05; not significant). The fertilization rate among excess oocytes in the P pattern group was 18% but was 39% (p < .0001) in the G pattern group. When an attempt was made to compensate for low morphology by increasing insemination concentration no significant difference in the pregnancy rate between the P and G pattern groups in the GIFT program was recorded. This was not the case when the in vitro fertilization rate was used as an endpoint. Insemination volume and swim-up concentration played no role in pregnancy rate.

Female↗
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Localization of wheat germ agglutinin lectin receptors on human sperm by fluorescence microscopy: utilization of different fixatives.

Wheat germ agglutinin (WGA) and Concanavalin A (Con A) seem to be the most widely studied of the lectins used to investigate sperm surface receptors. It has been reported that WGA receptors on human sperm membranes may be closely related to male fertility and that WGA receptor deficiency may cause human infertility. Although these claims may not be negated, there are many discrepancies in current literature regarding the localization and detection of human sperm WGA receptors. These discrepancies are probably the result of variations in the pretreatment of sperm before fixation and sperm-lectin interaction. This study investigated the effect of different fixation procedures on the localization of human sperm WGA receptors on normal spermatozoa. Comparative studies were performed on spermatozoa separated by wash-swim-up and fixed with 4% formaldehyde, 95% ethanol, or Bouin's solution, or nonfixed (air-dried) using fluorescein isothiocyanate-conjugated WGA. There were significant differences (p < .01) in the % WGA receptor localization following the different fixation procedures on samples receiving the same treatment previous to fixation. The different fixatives affect the plasma membrane to different degrees, resulting in an alteration of lectin receptor localization. To allow a better comparison and understanding of results, a standardized preparation procedure is recommended during studies using lectins as molecular probes. The results suggest than an aldehyde fixative may be the fixative of choice for sperm membrane studies.

Cell Membrane↗
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Comparison of motility characteristics and normal sperm morphology of human semen samples separated by percoll density gradient centrifugation.

The aim of this study was twofold: to investigate the ability of Percoll gradient centrifugation (52, 68, 84%) to fractionate semen samples according to motility quality and percentage normal morphology and to determine whether there is an association between sperm motility quality and percentage normal morphology. Sperm motility was evaluated using a Hamilton Thorn analyzer and normal sperm morphology was manually assessed according to the strict criteria (< or = 4, 5-14, and >14%). The majority of motility parameters and the percentage normal morphology were found to be significantly improved in the 84% Percoll fraction. The greatest effect was on the < or = 4% group, shifting the mean normal morphology percentage from 2.6 to 5.2%. Curvilinear velocity (VCL) and average path velocity (VAP) were the only two motility parameters that were significantly associated with the percentage normal morphology. Using a combined VCL, VAP vector the >14% group was found to have a significantly different value as compared to the 5-14 and < or = 4% groups. Percoll (discontinuous) gradient centrifugation can therefore play a significant role in the improvement of semen samples for use in assisted reproduction procedures. The VCL, VAP vector identified may also serve as an additional tool in the prediction of the fertility potential of sperm samples.

Analysis of Variance↗
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Slide preparation and staining procedures for reliable results using computerized morphology.

The purpose of this study was to standardize slide preparation and staining procedures to improve the efficiency and effectivity of the IVOS system on normal sperm morphology readings with regard to the strict criteria. Semen samples from patients attending the Reproductive Biology Unit, Tygerberg Hospital, were used. In experiment 1, five different Diff-Quik staining procedures, including the standard procedure, were evaluated on each of 22 patients and the effect of slide preparation within 1 h or more than 5 h after collection and the effect of immediate fixation versus fixation after 24 h were observed. In experiment 2, the manual evaluation time per slide (n = 20) by two technicians was compared with the time taken by computer. In experiment 1 the median % normal for the 5 different staining procedures was 6, 6.5, 9.5, 8.5, and 5.5%. No significant difference was found between the different staining procedures (p = .60, nonparametric Friedman test). In experiment 2 the mean time for manual assessment by two technicians was 3 min:6 s and 3 min:53 s per slide as compared to 4 min:39 s by computer. For experiment 1, slides can be prepared immediately or after 5 h. Fixation time also does not interfere with the computer's ability to identify normal forms. For experiment 2, the IVOS system is competitive regarding assessment time. Standardization of optimum staining procedures is important to ensure repeatability and comparability. Therefore, slides should be prepared immediately after liquefaction and fixed immediately after air drying.

Evaluation Studies as Topic↗
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Comparison between swim-up and glass wool column filtration of human semen in a gamete intrafallopian transfer program.

This study compared swim-up and glass wool filtration in both pregnancy outcome and fertilization of excess oocytes in patients undergoing gamete intrafallopian transfer. Gamete intrafallopian transfer patients were retrospectively included in the study group (n = 52). The criteria for inclusion were as follows: Semen had to have been prepared by means of glass wool filtration and at least 2 metaphase II oocytes had to have been transferred. Each patient from this group was then carefully matched with another patient according to specific criteria (number of metaphase II oocytes aspirated and transferred, normal sperm morphology, wife's age, the absence of anti-sperm antibodies, semen preparation by means of the swim-up procedure). Fourteen patients were matched with themselves (groups A1 and A2) and 38 patients were matched with another patient (groups B and C). The results indicate that there was no significant difference in the fertilization and pregnancy probabilities of sperm prepared by means of glass wool filtration or swim-up procedure. The comparative pregnancy rates for the groups were A1 (7.1%) versus A2 (7.1%) and B (28.9%) versus C (31.6%). Factors other than fertilization and pregnancy potential may have a greater influence on choosing the optimum sperm preparation procedure.

Female↗
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A comparative study using prepared and unprepared frozen semen for donor insemination.

The aim of this study was to compare the efficacy of pericervical insemination with unprepared semen With that of intrauterine insemination with prepared donor semen in a prospective, randomized, crossover clinical trial. Fifty-four subsequent patients who qualified for therapeutic donor insemination were randomized to receive alternately either an intrauterine insemination with thawed frozen donor semen, prepared by double wash and swim-up, or pericervical insemination with unprepared thawed frozen donor semen in subsequent treatment cycles, with each patient serving as her own control. Eleven pregnancies ensued from 54 cycles of intrauterine insemination (20.4% per cycle), and 2 pregnancies ensued from 58 cycles of pericervical insemination (3.4% per cycle, p = .005). Five pregnancies ensued during the first treatment cycle (13.2%), 4 during the second treatment cycle (17.4%), 2 during the third (13.2%), 1 during the fourth (7.1%), and 1 after the fourth treatment cycle (4.8%). The significantly better pregnancy rate from intrauterine insemination with prepared semen supports this treatment option. The decrease in pregnancy rate after the fourth treatment cycle confirms the need for alternative therapy after 4 failed cycles of therapeutic donor insemination.

Adolescent↗
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Effect of sperm preincubation time on GIFT pregnancy outcome.

This investigation was conducted to evaluate the effect of human sperm preincubation time on the pregnancy outcome in gamete intrafallopian tube transfers. This was determined in a retrospective study on gamete intrafallopian transfer patients (ideopathic infertility) using logistic regression with the covariates, preincubation time (< or = 60 and > 60 min) and age (< or = 35 and > 35 years). The study included 485 consecutive gamete intrafallopian transfer cycles in which 3 metaphase II oocytes were transferred. Pregnancy outcome was evaluated by beta hCG levels on days 12 and 16 and was confirmed by the presence of a fetal heart 8 weeks after the procedure by means of sonography. The wife's age and sperm preincubation time were significant covariates in the prediction of ongoing pregnancy. The odds ratio for age (< or = 35 years) was 2.2 in the prediction of ongoing pregnancy, while the age-adjusted odds ratios for preincubation time (< or = 60 min) was 1.9. There was a critical relationship between sperm preincubation time and GIFT success, which confirms the effect of the wife's age on pregnancy. Close coordination is of importance between the clinical scientist and the physician to restrict the preincubation time to 1 h. The sperm preincubation time is a significant factor that has to be considered in the prediction of ongoing pregnancy in GIFT patients.

Adult↗
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Significance of antisperm antibodies in female serum in a gamete intrafallopian transfer program.

The influence of antisperm antibodies in the female serum on fertilization and pregnancy rate in patients undergoing GIFT was assessed. A study group of 52 couples (69 cycles) with significant levels of antisperm antibodies in the female serum were compared to a control group of 749 couples (1185 cycles). Maternal serum or donor serum was used as growth medium supplement. The TAT test was performed for the detection of antisperm antibodies in the serum. Antisperm-antibody presence in female serum was associated with similar fertilization and pregnancy rates in a GIFT program compared to the control group. The type of serum used as growth medium supplement did not affect statistically the fertilization or pregnancy rate.

Adult↗
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Autosperm antibodies in treated, pregnant and non-pregnant immunologically infertile patients.

Twenty-two immunologically infertile couples (male partners had proven autosperm antibodies-positive mixed antiglobulin reaction test [MAR] and direct immunobead test [d-IBT]) were treated with washed spermatozoa used either in the gamete intrafallopian tube transfer (GIFT) or artificial insemination (AIH) procedures. Sixteen of the 22 couples (72.2%) fell pregnant with an ongoing pregnancy rate of 54.5% (12/22). The pregnant and non-pregnant groups were compared with regards to the sperm antibodies detected on the spermatozoa (MAR, d-IBT and sperm cervical mucus contact [SCMC] test) and in the serum and seminal plasma of the male partners (tray agglutination test [TAT], indirect immunobead test [i-IBT], and enzyme-linked immunosorbent assay [ELISA]). The semen parameters, motility, forward progression, count/ml and normal morphological forms were also compared. Statistical analysis showed no difference between the two groups (pregnant and non-pregnant) with regards to the antibody tests performed on semen, serum and seminal plasma. No difference was also seen between the semen parameters of the two groups. The washing of spermatozoa for the GIFT or AIH procedures may therefore be a successful method of treatment for immunologically infertile couples. The results also indicate no difference in the fertility prognosis for the two groups since antibody levels and semen quality were not different between the pregnant and nonpregnant group.

Antibodies, Anti-Idiotypic↗
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Comparative analysis of in vitro fertilization methods for establishing successful embryo transfer clinics.

A review of the laboratory methods and embryo transfer procedures used by seven of the foremost clinics in the world are discussed and evaluated in this article. Based on these facts an in vitro fertilization clinic was launched in 1983. In the second part of the paper the latest laboratory methods, i.e. ovulation induction plans and the embryo transfer technique of Tygerberg Hospital, are described. One hundred and thirty laparoscopies were performed since 1 May 1984-30 November 1984. Six hundred and one follicles were aspirated, 482 oocytes were obtained, 84% of all oocytes were fertilized, 112 patients reached the transfer stage and 28 pregnancies resulted. A clinical pregnancy rate of 21,4% per transfer and 18,5% per laparoscopy resulted.

Blood↗
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The technique for human embryo transfer at Tygerberg Hospital.

From 1 May 1984 to 30 April 1985 177 laparoscopies were performed; 684 oocytes were obtained and a fertilization rate of 85% per oocyte achieved. There was a pregnancy rate of 24.3% per embryo transfer (ET) and 20.9% per laparoscopy after 152 patients were subjected to ET. The ET technique at Tygerberg Hospital, using the Tomcat catheter, is described. An introducer--the FT introducer--designed to overcome technical difficulties in selected patients in passing the Tomcat catheter is described. Technical problems are easily overcome with this introducer. In 26 patients (17.1%) ET was performed using this instrument and 8 pregnancies resulted (a pregnancy rate of 30.7% per ET). This pregnancy rate indicates the value of the instrument.

Adult↗