Figure 7

CD4⁺T/CD8⁺T cell depletion ELISPOT assay. (1) CD4⁺T/CD8⁺T cell depletion. Two weeks after the DNA primary - 100 μg M1 intranasal boost immune strategy, the spleen lymphocytes were isolated and CD8 ⁺ T and CD4 ⁺ T cells were removed respectively by using MACs microbeads. (A) before the mixed lymphocyte sorting; (B) after removing the CD8⁺ T cells; (C) after removing the CD4⁺ T cells. (2) Collecting the MACs positive sorting cells. 2 × 10⁵ responding cells were incubated in 96-well PVDF plates coated with anti-IFN-γ monoclonal antibodies, the single peptide p22 was used to stimulate three kinds of different cells respectively, and the weakly positive single peptide p20 and p24 were used as the control (50 μg/ml peptide). (D) left-slash representing the whole spleen lymphocytes, cross hatch representing the spleen lymphocytes removing CD4⁺ T cells, right-slash representing the spleen lymphocytes removing CD8⁺ T cells. The values represent the averages of quadruplicate wells of 3 mice, and are expressed as means ± SD. The results were expressed as the number of SFC per 10⁶ input cells. *Significant differences compared to the mice in the whole splenocytes group (p < 0.05).