At 35 days, lymphocytes were isolated from the spleens as previously described. The cells were re-suspended at 5 × 106 cells/ml with RPMI Medium 1640 (Thermo Fisher Scientific) supplemented with 10% FBS, and then transferred into 96-well plates (100 μl per well), followed by PBS, Con A, B. subtilis-PRV, B. subtilis-gCa, and B. subtilis-gDa. The plates were incubated for 72 h at 37 °C. Then, the proliferative responses were detected using the standard CCK8 (Solarbio, China) method as previously described [32 (link)].
Free full text: Click here
