Antibody responses against avian reovirus nonstructural protein sigmaNS in experimentally virus-infected chickens monitored by a monoclonal antibody capture enzyme-linked immunosorbent assay.

Crude antigen preparations from avian reovirus (ARV)-infected chicken embryo fibroblasts (sigmaNS) or from bacterially expressed protein sigmaNS (esigmaNS) were captured by monoclonal antibody 1E1(MAb 1E1) against ARV nonstructural protein sigmaNS immobilized on the ELISA plates and were used as the MAb capture ELISA for antibody detection. Sixty one-week-old specific pathogenic free (SPF) chickens were divided into six groups and were vaccinated with live or inactivated ARV vaccine preparations in different combinations or inoculated with a virulent ARV strain. Sera collected from the birds were tested for their antibody responses to ARV nonstructural protein sigmaNS. Using the MAb capture ELISAs, the level of nonspecific binding reactions was tested on the serum samples obtained weekly from mock-infected SPF chickens from 1 to 25 weeks and compared to the results tested by the conventional ELISA. The results indicated that both MAb capture ELISAs had lower nonspecific bindings than those in the conventional ELISA, even in older birds. Antibody responses against ARV sigmaNS of the birds which received the inactivated vaccine twice (group I), inactivated vaccine followed by a live vaccine (group II), or a live vaccine followed by boosting with an inactivated vaccine (group III) were detected by MAb captured ELISA with sigmaNS crude antigens. The absorbance values increased rapidly at 1-2 weeks after boosting, approximated a peak at 5-6 weeks of age, and maintained this throughout the length of the experiment. The absorbance values of the MAb capture ELISA showed a good correlation to the SN titers ( r value > 0.85). On the other hand, serum samples from the birds which received the live vaccine twice (group IV) or were inoculated with a virulent ARV (group V) did not show antibody responses to sigmaNS, similar to those from the mock-infected birds (group VI), as the absorbance values maintained at a low level (below 0.5) throughout the length of the experiment. Similar results were obtained in the sera detected by MAb capture ELISA with crude esigmaNS antigens, except that the absorbance values in the sera from the birds in group III were gradually increased and later approximated a peak at 11 weeks of age and maintained this throughout the length of the experiments. The results suggest that MAb capture ELISAs can be readily used to detect antibody responses of the birds against ARV nonstructural protein sigmaNS which may reflect an immune status of a chicken flock, receiving ARV vaccine as long as including an inactivated vaccine.