Immunocytochemistry analysis of A549 cells labeling Eph receptor A2 with Rabbit anti-Eph receptor A2 antibody (HA723079) at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Eph receptor A2 antibody (HA723079) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

Eph receptor A2 was immunoprecipitated from 0.2 mg A549 cell lysate with HA723079 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA723079 at 1/1,000 dilution. Mouse Anti-Rabbit IgG kappa light chain secondary antibody (M1208-2) at 1/5,000 dilution was used for 1 hour at room temperature. Lane 1: A549 cell lysate (input) Lane 2: HA723079 IP in A549 cell lysate Lane 3: Rabbit IgG instead of HA723079 in A549 cell lysate Blocking/Dilution buffer: 5% NFDM/TBST Exposure time: 59 seconds; ECL: K1801