Western blot analysis of stray cat sera against Toxoplasma gondii and the diagnostic availability of monoclonal antibodies in sandwich-ELISA.

Abstract

A total of 198 sera from stray cats was assayed against Toxoplasma gondii antigen by western blot. Out of 198 sera assayed, 26 sera (13.1%) showed typical blot patterns against T. gondii. When spotted by ELISA absorbance and indirect latex agglutination test (ILAT) titer, all 26 cases were distributed over the cut-off value of ELISA whereas 24 cases (92.3%) were in the positive range of 1:32 or higher and 2 cases in negative range by ILAT. Among western blot negative 172 sera, 162 cases were negative in both ILAT and ELISA while 10 cases were reactive falsely such that three cases were ILAT positive with 1:32 titer and 9 cases were ELISA positive (2 cases overlapped). These 10 cases reacted peculiarly without typical binding pattern in Western blot. Sandwich-ELISA was performed with monoclonal antibodies (mAbs) of Tg563 (30 kDa, SAG1), Tg505 (22 kDa, SAG2), Tg605 (43 kDa, SAG3), Tg556 (28 kDa. GRA2), Tg737 (32 kDa, GRA6), Tg695 (66 kDa, ROP2), Tg786 (42 kDa, ROP6), and Tg621 (32 kDa, anonymous but cytosolic) clone, respectively. All western blot-positive cases were in the positive range and negative cases in the negative range clearly. Among the 10 false reactive cases, 3 cases were in the positive range with one or more mAbs. All mAbs used in this study were confirmed to be specific to T. gondii infection as a standardized sandwich-ELISA to differentiate it from other pathogens.