Sample:
Lane 1: Mouse Spleen tissue lysates
Lane 2: Mouse Thymus tissue lysates
Lane 3: Mouse Lymph node tissue lysates
Lane 4: Mouse Lung tissue lysates
Lane 5: Rat Spleen tissue lysates
Lane 6: Rat Thymus tissue lysates
Lane 7: Human K562 cell lysates
Lane 8: Human U-2 OS cell lysates
Lane 9: Human U-87 MG cell lysates
Primary: Anti-IL-7Ra (SL1540R) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 50 kD
Observed band size: 70 kD
Blank control: U937(blue).
Primary Antibody: Rabbit Anti-IL-7Ra antibody(SL1540R), Dilution: 1μg in 100 μL 1X PBS containing 0.5% BSA;
Isotype Control Antibody: Rabbit IgG (orange) ,used under the same conditions.
Secondary Antibody: Goat anti-rabbit IgG-PE(white blue), Dilution: 1:200 in 1 X PBS containing 0.5% BSA.
Protocol
The cells were fixed with 2% paraformaldehyde (10 min).Primary antibody (SL1540R, 1μg /1x10^6 cells) were incubated for 30 min on the ice, followed by 1 X PBS containing 0.5% BSA + 10% goat serum (15 min) to block non-specific protein-protein interactions. Then the Goat Anti-rabbit IgG/PE antibody was added into the blocking buffer mentioned above to react with the primary antibody at 1/200 dilution for 30 min on ice. Acquisition of 20,000 events was performed.