Implementation of immunohistochemical methods in diagnostics of bovine sarcocystosis

Abstract

Ispitivanje sarkocistioze goveda obavljeno je na tri starosne kategorije goveda primenom direktnih i imunohistohemijskih metoda. Primenjene su direktne metode kompresije, digestija tripsinom i histološka metoda, a od imunohistohemijskih metoda korišćene su peroksidaza anti-peroksidaza (PAP),visoko senzitivna streptavidin biotin imunoperoksidaza kompleksna tehnika (LSAB+) i indirektna imunofluorescencija (IF). Primarni antiserum kunića dobijen na zoite Sarcocystis cruzi syn. S. bovicanis korišćen je za sva imunohemijska i imunohistohemijska istraživanja.Za određivanjeunakrsnereaktivnostikorišćenje kao primarni antiserum kunića na antigene zoita S. ovifelis. Oba primarna antiseruma proizvedena su u laboratoriji, zbog nedostatka komercijalnih preparata na tržištu. Procena osetljivosti, specifičnosti i efikasnosti imunohistohemijskih metoda izvedena je na bazi poredenja sa direktnim metodama: kompresije, digestije tripsinom i histološkom metodom. Primenjene imunohistohemijskeanalizesu vrlospecifičneu dokazivanjusarkocistiozekod goveda, posebno LSAB*tehnika,a po osetljivosti u otkrivanju ove infekcije slične su metodi digestije tripsinom. Dobijeni antiserum kunića na antigenezoita S. bovicanis daoje dobre rezultate u dijagnostikovanju homologne vrste. Dokazana unakrsna reaktivnost homolognih i heterolognih antiseruma i antigena, kao imunohemijska reakcija potpunog identiteta ujednom delu antigene strukture, ustanovljenaje kod primarnog antiserumakunića na zoite S. ovifelis, što ukazujeda se i heterologni antiserumi mogu da koriste u imunohistohemijskim reakcijama dijagnostikovanja sarkocistioze goveda.

Bovine sarcocystosis was examined in three age categories of cattle using direct and immunohistochemical methods. The direct methods of compression, trypsin digestion and a histological method were applied, and the applied immunohistochemical methods included peroxidase anti-peroxidase (PAP), the highly sensitive streptavidin biotin immunoperoxidase complex technique (LSAB+) and indirect immunofluorescence (IF). Primary rabbit antiserum obtained for the zoites Sarcocystis cruzi syn. S. bovicanis was used for all immunochemical and immunohistochemical examinations. Cross reactivity was determind using primary rabbit antiserum to antigens of S. ovifelis zoites. Both primary antiserums were produced in the laboratory, as no commercial preparations were available on the market. The evaluation of the sensitivity and efficacy of the immunohistochemical methods was performed on the grounds of comparisons With the direct methods: compression, trypsin digestion and a histological method. The applied immunohistochemical analyses are very specific in proving sarcocystosis in cattle, especially the LSAB+ technique, and, according to their sensitivity in detecting this infection, they are similar to the method of trypsin digestion. The obtained rabbit antiserum against antigens of S. bovicanis zoites yielded good results in the diagnosis of the homologous specie. Proven cross reactivity of the homologous and heterologous antiserums and antigens, as an immunochemical reaction of complete identity in one part to the antigen structure, was established in primary rabbit antiserum to S. ovifelis zoites, which indicates that heterologous antiserums can also be used in immunohistochemical reactions in the diagnostics of bovine sarcocystosis