B Cell Activation on an Antigen-Coated Coverslip

Prepare the antigen solution by adding anti-BCR and B220 in PBS to a final concentration of 10 micrograms per mL and 7.5 micrograms per mL respectively. Next, set the coverslip over a lid of 24-well plate covered with paraffin film, and add 40 microliters of antigen solution. Seal the plate, and then incubate at four degrees overnight.

To start the activation in coverslip, first, wash them with PBS, and let them dry for a few minutes. Next, add 150,000 B cells, and incubate at 37 degrees for the different time points. In both cases, when activating either with beads or antigen-coated slides, we recommend starting with the longest time point, and then proceeding to the shorter ones.