Biomedical subjects

P F Levay

Publications and source records attributed to P F Levay.

6 recordsLinked to original sources

PubMed1997

The detection of blood contamination in human follicular fluid.

PURPOSE: The aim of this study was to compare the reliability of the methods conventionally used to identify low levels of blood contamination in human follicular fluid (hFF) as applicable in the clinical environment. METHODS: Follicular fluid (n = 339) and plasma samples (n = 20) were collected from patients (n = 138) attending the Centre for Fertility Studies, HF Verwoerd Hospital, University of Pretoria, South Africa. hFF blood contamination was assessed by means of (a) visual inspection, (b) hematocrit (Hct), (c) spectrophotometric analysis, (d) spectrophotometric hemoglobin kit, and (e) Combur-9-test urine sticks. RESULTS: (1) Neither hematocrit nor spectrophotometry provided reliable detection at low levels of blood contamination. (2) Visual inspection presented with a better discriminatory ability than either Hct or spectrophotometry. (3) Combur-9-test sticks identified up to 50% of blood-contaminated fluids. (4) Spectrophotometrically determined hemoglobin levels presented with weak discriminatory abilities for detecting blood-contaminated fluids. CONCLUSIONS: Visual inspection as performed in this study provides a fast and relatively reliable method for the determination of blood-contaminated hFFs. In a laboratory environment, however, it would be recommended that a combination of visual inspection, Hct, and spectrophotometric evaluation be employed for the selection of blood-free fluids.

Blood↗

PubMed1995

The effect of seminal plasma on human sperm-zona pellucida binding.

The aim of this study was to determine the effect of different concentrations of seminal plasma (SP) in insemination medium on human sperm-zona pellucida (ZP) binding. The effects of fresh SP [50% (n = 10) and 1% (n = 9)], frozen-thawed versus fresh SP [5% (n = 10)] and frozen-thawed SP [5% (n = 10) and 1% (n = 9)] on sperm-ZP binding were examined using hemizona assays. The validity of the hemizona assay as performed in this study was also examined. Relative sperm binding percentages were determined for respective hemizonae, and the hemizona index was calculated for each experimental group and compared statistically. The 50% native SP inhibited sperm-ZP binding by approximately 70% (P = 0.0051), while 1% native SP enhanced sperm-ZP binding by approximately 350% (P = 0.0051). Significantly more spermatozoa (mean +/- SD, 172.00 +/- 54.12) bound to zonae in the presence of 5% frozen-thawed SP than with 5% SP that had not been frozen (mean +/- SD, 127.00 +/- 69.18; P = 0.0431). The 1 and 5% frozen-thawed SP stimulated sperm-ZP binding by approximately 400 and approximately 250% respectively (P = 0.0077 and 0.0051 respectively). It is concluded that SP concentrations found in insemination media during assisted reproductive techniques do not inhibit but in fact enhance sperm-ZP binding.

Cadaver↗

PubMed1995

Effectiveness of various sperm processing methods in removing seminal plasma from insemination media.

The effectiveness of the wash and swim-up (1 and 2 wash cycle), two layer Percoll gradient, SpermPrep and underlay sperm preparation methods in removing seminal plasma from insemination suspensions was investigated. The number of wash cycles needed to rid sperm suspensions of seminal plasma (n = 15) was also determined. All sperm preparation methods were compared to the control wash and swim-up method, performed using Earle's buffered salt solution (EBSS), without serum supplementation so as not to mask seminal plasma concentrations. Control processing comprised 1 ml semen subjected to two wash cycles in EBSS followed by a swim-up period of 60 min under 5% CO2 in air. The Percoll method comprised 1 ml semen layered on a discontinuous 36 and 81% Percoll gradient (n = 14). In the SpermPrep method, 1 ml semen was run through a SpermPrep II column (n = 10), and underlay samples (n = 12) were processed by two wash cycles, after which sperm pellets were resuspended in the remaining medium, layered under 1.2 ml EBSS and allowed to swim up under 5% CO2 in air for 1 h. Seminal plasma and supernatant fractions obtained after each processing phase were stored at -20 degrees C and protein concentrations were determined by spectrophotometry. The wash and swim-up method was the most effective in removing seminal plasma from sperm suspensions, followed by the two-layer Percoll gradient, underlay and finally the SpermPrep II processing methods.

Chemical Fractionation↗

PubMed1994

Interleukin-1 beta, interleukin-6, and growth hormone levels in human follicular fluid.

PURPOSE: To investigate possible relationships of interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), and growth hormone (GH) with biochemical variables in human follicular fluid (FF) and selected in vitro fertilization (IVF) parameters. METHODS: A total of 67 FF samples (n = 67 patients undergoing oocyte retrieval for IVF) was evaluated. IL-1 beta, IL-6, GH, hLH, FSH, PRL, hCG, testosterone, total protein, fibrinogen, sialic acid, alpha 1-antitrypsin, plasminogen levels, and spectrophotometric absorbance at 458 nm were analyzed for selected FF. IL-6 and GH levels of serum and FF samples were also compared (n = 23). RESULTS: Immunoreactive levels of IL-1 beta, IL-6, and GH were detected in all FF samples. A positive correlation existed for IL-6 (r = 0.5069, P = 0.0161) when serum-to-FF levels were compared (concentration ratio, 1:1.857). Smaller-volume follicles (< 4 ml) were associated with high IL-1 beta levels (P = 0.0229), and an additional tendency of IL-1 beta to decrease with increasing embryo cleavage and scoring was observed. With the exception of a weak positive correlation between follicular IL-1 beta and testosterone levels (r = 0.3128, P = 0.025), no other relationship with biochemical variables or IVF parameters (etiology, e.g., endometriosis) could be implicated. CONCLUSIONS: Substantially higher IL-6 levels occurred in FF compared to serum, thus supporting intrafollicular production. Interleukin-1 beta, IL-6, and GH levels in FF are, however, unsuitable markers for in vitro fertilization outcome.

Adult↗

PubMed1994

Electron microscopic study of human sperm membrane isolation.

OBJECT: Our purpose was to isolated pure, homogeneous human sperm membranes, free of cellular contaminants. METHODS: Donor semen samples collected after masturbation were stored at -70 degrees C and eventually pooled. Each attempt at sperm membrane isolation required 800 x 10(6) spermatozoa which were sonicated by ultrasound (40% output; Vibra Cell). The effect of sonication time (3 x 5, 3 x 15, and 180 sec) on membrane isolation was investigated. Sonicated samples were centrifuged (500 g, 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 (100,000 g, 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 x 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. CONCLUSION: The advantage of this method is its simplicity. The drawback of this method is the large number of spermatozoa required for membrane purification.

Cell Fractionation↗

PubMedDate not supplied

Lactoferrin: a general review.

Lactoferrin is a 703-amino acid glycoprotein originally isolated from milk. Plasma lactoferrin is predominantly neutrophil derived but indications are that it may also be produced by other cells. Lactoferrin in body fluids is found in the iron-free form, the monoferric form and in the diferric form. Three isoforms of lactoferrin have been isolated, ie two with RNase activity (lactoferrin-beta and lactoferrin-gamma) and one without RNase activity (lactoferrin-alpha). Receptors for lactoferrin can be found on intestinal tissue, monocytes/macrophages, neutrophils, lymphocytes, platelets, and on certain bacteria. A wide spectrum of functions are ascribed to lactoferrin. These range from a role in the control of iron availability to immune modulation. More research is necessary however to obtain clarity with regard to the exact mechanism of action of lactoferrin.

Lactoferrin↗