OBJECTIVETo discuss the enzyme linked immune spot test (ELISPOT) detected the cellular immune response induced by human Bocavirus (HBoV) VP2 virus-like particles (VLPs).

METHODSAfter immunized by HBoV VP2 VLPs, the specific cellular immune response in mice were detected by ELISPOT assay, observe the ELISPOT results at the conditions of different polypeptide stimulate, different cell culture time, different cell concentration and different specific stimulus peptide concentration, then screening the right ELISPOT experimental conditions and establish the ELISPOT method.

RESULTSThe spots induced by HBoV1 VLPs immunized mice spleen lymphocytes stimulate with polypeptide P3 (GYIPIENEL) and P5 (LYQMPFFLL)were 233 spots/10(6) cells and 157 spots/10(6) cells,spots induced by HBoV2 VLPs immunized mice spleen lymphocytes stimulate with polypeptide P8 (GYIPVIHEL) were 113 spots/10(6) cells; 24 hours is the best time for culture, at this time HBoV1 and HBoV2 groups specificity secretion IFN-gamma ratio were 232 spots/10(6) cells and 119/10(6) cells; Best concentration of mice spleen lymphocyte is 5 x 10(5), right now HBoV1 and HBoV2 group specificity secretion IFN-gamma ratio were 232 spots/10(6) cells and 108/10(6) cells; Best concentration of polypeptides is 10 microg/ml, HBoV1 and HBoV2 group specificity secretion IFN -gamma ratio were 233 spots/10(6) cells and 96/10(6) cells.

CONCLUSIONSHBoV1 and HBoV2 specificT-cell epitope in BABL/c mice were P3, P5 (HBoV1) and P8 (HBoV2). The best experiment condition were: cell cultivated for 24 h, cells concentration for 5 x 10(5) cells/well, stimulating polyperides concentration for 10 microg/ml, it can use to study the cellular immune induced by HBoV in mice.