Bilateral endoscopic totally extraperitoneal (TEP) inguinal hernia repair does not impair male fertility.

PURPOSE: Endoscopic totally extraperitoneal (TEP) hernia repair with polypropylene mesh has become a well-established technique. However, since the mesh is placed in close contact with the spermatic cord, mesh-induced inflammation may affect its structures, possibly resulting in impaired fertility. The aim of this observational prospective cohort study was to assess fertility after bilateral endoscopic TEP inguinal hernia repair in male patients. METHODS: Fifty-seven male patients (22-60 years old) with primary, reducible, bilateral inguinal hernias underwent elective bilateral endoscopic TEP hernia repair with use of polypropylene mesh. The primary outcome was testicular perfusion; secondary outcomes were testicular volume, endocrinological status, and semen quality. All patients were assessed preoperatively and 6 months postoperatively. RESULTS: Follow-up was completed in 44 patients. No statistically significant differences in measurements of testicular blood flow parameters or testicular volume were found. Postoperative LH levels were significantly higher [preoperative median 4.3 IU/L (IQR 3.4-5.3) versus postoperative median 5.0 IU/L (IQR 3.6-6.5), p = 0.03]. Levels of inhibin B were significantly lower postoperatively [preoperative median 139.0 ng/L (IQR 106.5-183.0) versus postoperative median 27.0 ng/L (IQR 88.3-170.9), p = 0.01]. No significant changes in FSH or testosterone levels were observed. There were no differences in semen quality. CONCLUSIONS: Our data suggest that bilateral endoscopic TEP hernia repair with polypropylene mesh does not impair fertility, as no differences in testicular blood flow, testicular volume, or semen quality were observed. Postoperative levels of LH and inhibin B differed significantly from preoperative measurements, yet no clinical relevance could be ascribed to these findings.

Rapid in vitro micro-cytotoxicity tests for the detection and quantitation of neutralizing antibodies to both viruses and toxins.

A generally applicable method was developed for the rapid and quantitative detection of both toxin- and virus neutralizing antibodies. The method was optimized for three different biological agents, i.e., Shigella toxin, influenza viruses (A/Beying, A/Taiwan and B/Yamagata) and Chikungunya virus. The in vitro micro-cytotoxicity tests developed for the detection and quantitation of neutralizing antibodies are based on the inhibition of the virus- or toxin-induced cytotoxic effect by antibodies. As a result of the cytotoxicity, infected cells are no longer attached to the solid phase and can be easily removed. Thereafter, the proteins of the remaining living cells are stained. After removing the excess dye, the remaining dye is dissolved and the absorbance values are measured. The neutralization titers are determined from the absorbance values. Since the tests are performed in wells of microtiter plates, the in vitro micro-cytotoxicity tests are less laborious and consume less reagent in comparison with classical neutralization tests.

Automated high-performance liquid chromatographic determination of chloramphenicol in milk and swine muscle tissue using on-line immunoaffinity sample clean-up.

An on-line high-performance liquid immunoaffinity chromatographic (HPLIAC) system for the direct determination of chloramphenicol in milk and swine muscle tissue is described. The system consisted of a dual-column system in which an HPLIAC column was directly coupled to an RP-8 high-performance liquid chromatographic column. Skimmed and deproteinated milk or aqueous muscle tissue extract was directly injected into the HPLIAC column. After a washing step with phosphate-buffered saline, chloramphenicol was desorbed by a glycine-NaCl buffer (pH 2.8) and directly concentrated on the RP-8 column. Next, chromatography was carried out using acetonitrile-sodium acetate buffer as the mobile phase. Chloramphenicol was detected at 280 nm. Mean recoveries from spiked raw milk were 70 +/- 2% (1-50 micrograms/kg) and from spiked swine muscle tissue 64 +/- 2% (10-50 micrograms/kg). The calibration curves were linear in the range 1-200 micrograms/kg spiking levels. Limits of determination were 1 microgram/kg for milk and 10 micrograms/kg for muscle tissue.

Analysis of chloramphenicol residues in swine tissues and milk: comparative study using different screening and quantitative methods.

Both screening and quantitative methods for chloramphenicol residues in swine tissues and milk were compared, using samples from animals treated with chloramphenicol. For screening purposes a previously developed streptavidin-biotin enzyme-linked immunosorbent assay and a commercially available immunochemical card test were used. For quantitative purposes two previously developed high-performance liquid chromatographic procedures were applied using antibody-mediated clean-up and solid-phase extraction. Some improvements in both methods were also described. The results obtained with the screening tests and those obtained with the quantitative methods correspond well with each other. Using a combination of these methods, an effective control of residues of chloramphenicol can be performed in milk from the 1 microgram/kg level and in swine tissues from the 10 micrograms/kg level.

Sensitive streptavidin-biotin enzyme-linked immunosorbent assay for rapid screening of chloramphenicol residues in swine muscle tissue.

A sensitive, competitive enzyme-linked immunosorbent assay (ELISA) for chloramphenicol (CAP) in swine muscle tissue has been developed. The ELISA is based on an earlier procedure. To improve sensitivity, different optimization procedures were investigated. The introduction of a streptavidin-biotin system and the use of a coating antigen with a lower CAP incorporation resulted in the most sensitive ELISA: the CAP concentration giving 50% inhibition decreased from 125 ng/mL to 3.0 ng/mL. This ELISA procedure was applied for a rapid screening of CAP residues in swine muscle tissue. The tissues were extracted with demineralized water. A concentrated phosphate-buffered saline solution was added to the filtered aqueous extract and this sample solution was directly submitted to the ELISA procedure. The results were compared to values obtained by analysis of a corresponding blank. This blank was prepared by treating a part of the aqueous sample solution with an immobilized monoclonal antibody preparation. This treatment was necessary because aqueous extracts of different swine muscle tissues showed a high variation in dose-response curves, probably caused by the complexity and variability of the matrix. In spiked tissues, the presence of CAP at concentrations of 10 micrograms/kg and higher can be easily demonstrated.

Monoclonal antibody-mediated clean-up procedure for the high-performance liquid chromatographic analysis of chloramphenicol in milk and eggs.

A simple, rapid and specific sample preparation method based on antibody-mediated clean-up for the determination of chloramphenicol (CAP) in milk and eggs was developed. Skimmed milk and centrifuged egg homogenates were filtered and directly applied to immunoaffinity columns which were prepared by coupling monoclonal antibodies against CAP to a carbonyldiimidazole-activated support. Using a 0.2 M glycine, 0.5 M NaCl (pH 2.8) solution as an eluent, the immunoaffinity columns can be used more than 30 times without a decrease in column capacity. In subsequent high-performance liquid chromatographic analysis, no matrix interferences were observed. Good recoveries were obtained at spiking levels of 1-100 micrograms kg-1. Due to the high specificity of the clean-up procedure, the limit of detection can be lowered by increasing the test portion. Concerning milk, the limit of detection was successfully lowered to 20 ng kg-1 by increasing the test portion to 11 (recovery 99%). The method was applied to eggs produced by hens treated with CAP. The results are compared with those obtained by solid-phase extraction using silica gel.

Determination of chloramphenicol in swine muscle tissue using a monoclonal antibody-mediated clean-up procedure.

A rapid and specific clean-up procedure based on immunoaffinity chromatography for the high-performance liquid chromatographic determination of chloramphenicol (CAP) in swine muscle tissue at the 10 micrograms kg-1 level is described. For preparation of the columns, monoclonal antibodies against CAP were coupled to cyanogen bromide-activated Sepharose. After immunosorption of CAP from the aqueous meat extract, CAP was eluted with methanol. Mean recoveries from spiked swine muscle tissue were 70 +/- 6% (10-50 micrograms kg-1 spiking levels). The loss of CAP can be completely attributed to the extraction; no loss was observed during the antibody-mediated clean-up.

An enzyme-linked immunosorbent assay for the determination of chloramphenicol using a monoclonal antibody. Application to residues in swine muscle tissue.

A competitive enzyme-linked immunosorbent assay (ELISA) using a monoclonal antibody is described for the determination of chloramphenicol (CAP) residues in swine muscle tissue. The limit of detection of standard solutions is established to be 25 ng ml-1 = 2.0 ng CAP per well). The very high specificity of the monoclonal antibody for CAP is expressed by the insignificant cross-reactivity with other antimicrobial agents and with structurally related compounds. By means of the rapid sample preparation method described earlier for the CAP determination using high-performance liquid chromatography (HPLC), residue levels of CAP in swine muscle tissue above 5 micrograms kg-1 can be easily quantitated. The muscle samples show good recovery percentages at 10-50 micrograms kg-1 spiking levels. The results obtained by the ELISA method were confirmed by HPLC.