Produção de antígenos para diagnóstico da artrite encefalite caprina empregando cultivo de células CorFC com baixo teor de soro fetal bovino

Abstract

The objective of this work is to develop serological tests (AGID and ELISA -ELISAi) using antigens of the virus Arthritis Encephalitis Goat (CAEV) obtained from a goat corneal cell line (CorFC cells) grown with 0.1% FBS. For AGID antigens were obtained from cells cultured in MEM, DMEM / F12 and RPMI 1640. Of the 163 tested sera samples (28 positive and 135 negative), 28 (17.17%) were positive for MEM and Ag Ag DMEM / 12 and 29 (17.79%) to Ag is RPMI 1640. Comparing the results obtained with three Ag was observed almost perfect agreement kappa adjusted (0.98 to 1.00). The Ag RPMI 1640 antigen, which showed better quality of precipitation lines in AGID and best performance was evaluated in comparison to a commercial Ag (Kit for the diagnosis of CAE - AGID; Unika biotech, Recife). The test of 797 sera with commercial Ag showed 216 positive and 581 negative; RPMI 1640 with Ag were observed 225 positive and 572 negative, corresponding to sensitivity (Se) and specificity (Es) for 100% and 98.45%, respectively. Globally, there has been adjusted (kappa = 0.97) between the almost perfect results with both Ag. For ELISAi empregandos native viral antigens were obtained by a simple process (clarification, dialysis and the treatment with triton X-100) to from cells cultured in RPMI 1640. After standardization were made the ELISAi performance evaluations. To assess the repeatability of the observed ELISAi RPMI CV entreplacas 4.4 and 17.3 and between 18.4 and 5.1 days for negative and positive sera. The results of 722 sera tested with the kits to diagnose LVPR AGID / commercial ELISAi (Unika biotech, Recife) showed 166 positive and 556 negative; with RPMI were observed ELISAi 171 positive and 551 negative, and if Sp corresponding to about 97.59% and 98.38%, respectively, with agreement adjusted kappa 0.95, considered almost perfect. These results demonstrate that one can establish an AGID an ELISA for highly sensitive and specific diagnosis of LVPR using a simplified model for obtaining of antigens, employing CorFC cells with low levels of FBS. Depending on the cell type and medium used can be produced LVPR antigens with low FCS content, which agrees with the animal welfare, represents a reduction of input costs and protein purification processes.