Characterization of microRNA in the follicular fluid of patients undergoing assisted reproductive technology.

Female fertility plays a decisive role in the reproduction of mammals, with related issues that include oocyte or embryo quality, establishment of pregnancy, and the physiology of the tissues that contribute to reproduction and metabolic disorders associated with reproductive failure. Although reproductive failure may be attributed to various factors in different species, female infertility is largely controlled by a number of molecular signals that can be regulated in a cycle- and tissue-dependent manner.

Profiling bovine blastocyst microRNAs using deep sequencing.

MicroRNAs (miRNAs) are known to control several reproductive functions, including oocyte maturation, implantation and early embryonic development. Recent advances in deep sequencing have allowed the analysis of all miRNAs of a sample. However, when working with embryos, due to the low RNA content, miRNA profiling is challenging because of the relatively large amount of total RNA required for library preparation protocols. In the present study we compared three different procedures for RNA extraction and prepared libraries using pools of 30 bovine blastocysts. In total, 14 of the 15 most abundantly expressed miRNAs were common to all three procedures. Furthermore, using miRDeep discovery and annotation software (Max Delbrück Center), we identified 1363 miRNA sequences, of which bta-miR-10b and bta-miR-378 were the most abundant. Most of the 179 genes identified as experimentally validated (86.6%) or predicted targets (13.4%) were associated with cancer canonical pathways. We conclude that reliable analysis of bovine blastocyst miRNAs can be achieved using the procedures described herein. The repeatability of the results across different procedures and independent replicates, as well as their consistency with results obtained in other species, support the biological relevance of these miRNAs and of the gene pathways they modulate in early embryogenesis.

Complement fixation reaction in toxoplasmosis.

In toxoplasmosis serodiagnosis the complement fixation reaction (CF) is barely sensitive and specific and supplies results below the general standard levels of this technique. The reasons for this deficiency are discussed and detected in the preparation modalities of the toxoplasma antigens. Two diagnostic antigens are prepared and evaluated; the former, a suspension of whole toxoplasma (WT) the latter a total extract of toxoplasma (TET). The antigens are characterized by the preparative process, culture host and the extractive technique with a modified ultrasonic disintegrator. The antigens are used in the CF reaction, performed with the modified LBCF method, with a 100% hemolysis reading (H 100). The LBCF-H 100 reaction with WT and TET antigens is evaluated parellely to the indirect immunofluorescence reaction (IF) and dye test (DT) on 2514 human sera from cases os suspected toxoplasmosis and pregnant women. The analysis of the serological results pointed out that the LBCF-H 100 reaction performed with WT antigen, shows a sensitivity and specificity equivalent to that of the DT. The LBCF-H 100 reaction with TET antigen extends the range of the antibodies detectable with WT antigen. The optimal serological combination to identify the highest number of seropositive cases of toxplasma infection is given by LBCF-TET and IF reactions.

Purification and concentration of influenza inactivated viruses by continuous-flow zonal centrifugation.

A mathod is described for the purification, on an industrial scale, of influenza viruses grown in allantoic cavity of embryonated eggs. The mehtod consists of combining continuous-flow centrifugation with zonal centrifugation in a sucrose (36.6 per cent-52.5 per cent w/v) density gradient. The sample flow rate is approximately 3.7 litres/h and the volumes treated vary between 3 and 33 litres of allantoic fluid. Both the recovery of the virus and the degree of concentration and purification result satisfactory.