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M. Kolesárová, R. Herich, M. Levkut, J. Čurlík and M. Levkut

Abstract

PCR amplification of specific DNA regions is a powerful tool for retrospective studies, but not all preservation or fixation methods render DNA that is suitable for subsequent amplification. Several factors affect sensitivity of polymerase chain reaction (PCR) amplification. There were reported the effects of commonly used fixation solutions — 10 % neutral buffered formalin, 20 % neutral buffered formalin and Carnoy’s solution and the efficiency of PCR amplification in fresh tissue and paraffin (or wax) embedded samples of Cysticercus ovis. DNA from samples was isolated and PCR product of 1300 bp was amplified. Results indicated that the samples fixed in Carnoy’s solution produced reliable amplification of desired fragments. The samples that were fixed in 10 % and 20 % neutral buffered formalin brought negative results.

Open access

V. Revajová, M. Levkut, A. Aldawek, R. Herich, E. Dvorožňáková and I. Krupicer

Abstract

To estimate the effect of lambs’ immunoreactivity to multiple Toxocara infection, subpopulations of lymphocytes, determination of specific IgG levels, proliferation activity of splenic lymphocytes, and metabolic activity of granulocytes were performed. Five-month-old Valaška lambs were daily infected with 1000 embryonated Toxocara canis eggs for 23 days and immune responses were studied up to 49 days post infection (dpi). The number of leukocytes and neutrophiles was no significantly higher on 12 hrs pi in infected group. Absolute counts of eosinophiles were increased on 14 dpi and significantly increased on 21 d in infected animals. Greater numbers of CD2+ T lymphocytes were observed during the course of the experiment in the peripheral blood of infected animals. The level of CD4+ cells was lower 42 dpi, but the number of CD8+ cells was higher in the experimental animals. IgM positive B cells were significantly increased on 28 dpi, and monocytes on 12 hours, 14 and 35 dpi in infected animals. Production of anti-T. canis IgG was significantly enhanced on 28 dpi in infected lambs. Significant increased proliferative response of splenic T and B lymphocytes was found on 14 and 21 dpi. Metabolic burst of granulocytes demonstrated the decrease of percentage values from 3 dpi. Larvae of T. canis were microscopically observed in the peripheral blood from 14 to 35 dpi. The multiple reinfection of sheep with 1000 T. canis eggs caused eosinophilia, increased the proliferation activity of B-and T-cells and increased the production of T. canis specific antibodies. The cooperation of immune cells has directed to kill the larvae in the peripheral blood and trapping them in granulomas.