The expressed EdIII, not the NusA -Tag protein, was detected by antibodies that detect the E proteins of the tick-borne flavivirus by Western blot. These

results indicated that EdIII can be useful as the antigen in the diagnosis ELISA. One hundred and twenty serum samples from wild rodents captured in Kamiiso, Hokkaido, were tested for TBE virus-specific antibodies by EdIII-ELISA, SP-ELISA and the neutralization test. The detection accuracy of each ELISA was evaluated by comparing the results between the neutralization test and the ELISAs. Figure 2 shows the sensitivity and specificity of the EdIII-ELISA by comparison with the neutralization test, using the corresponding cut-off values. The sensitivity of the EdIII-ELISA decreased with increasing cut-off values, while the specificity increased. The difference between the sensitivity and MK-2206 chemical structure specificity was a minimum selleck screening library value when a cut-off value of 0.61 was used. Then at a cut-off value of 0.64, a higher specificity (80.0%, 68/85) and equal sensitivity (77.1%, 27/35) were obtained, compared to the cut-off value of 0.61 (Table 1). The SPs were expressed by the transfection of the plasmid pCAGprME into 293T cells

and precipitated using PEG solution as described previously (15). Anti-E protein rabbit IgG was prepared by immunization of a rabbit with the EdIII in order to use it as the capture antibody in the SP-ELISA (23). The anti-E protein rabbit IgG was confirmed to be reactive to both the E protein from the authentic

TBE virus antigen and the SPs (Fig. 3). These results indicated that the anti-E protein rabbit IgG can be useful for the capture antibody of the diagnostic SP-ELISA. Figure 4 shows the sensitivity and specificity of the SP-ELISA by comparison with the neutralization test, using the corresponding cut-off values. The sensitivity of the SP-ELISA decreased with increasing cut-off values, while the specificity increased. The difference between the sensitivity and specificity was at a minimum value when a cut-off value of 0.042 was used. Then at a cut-off value of 0.089, a higher specificity (100%, 85/85) and equal sensitivity (91.4%, 32/35) were obtained, compared to the cut-off value of 0.042 (Table 2). To investigate Bumetanide whether our ELISAs using recombinant antigens can be applied to the epizootiological survey, wild rodent samples were collected in Khavarovsk, Russia, an area in which many TBE patients were reported (24), and examined for anti-TBE virus antibodies by the ELISAs. Twenty-nine serum samples from wild rodents were tested by the EdIII-ELISA and the SP-ELISA, and the same three samples were diagnosed as positive by both ELISAs (Table 3). The three samples were also positive for the neutralization test and the other 25 samples, which were negative for the ELISAs, were also negative for the neutralization test.