Anti-PD-1 [J43]

Binding curve of anti-PD-1 antibody J43 (orb758881) to recombinant mouse PD-1 Fc-Fusion Protein. ELISA Plate coated with recombinant mouse PD-1 Fc-Fusion Protein at a concentration of 5 µg/ml. A 3-fold serial dilution from 10000 ng/ml was performed using orb758881. For detection, a 1:4000 dilution of HRP-labelled anti-rabbit antibody was used.

Flow cytometry using the Anti-PD-1 antibody J43. Paraformaldehyde fixed mouse splenocytes permeabilized with 0.5% Triton were stained with anti-unknown specificity antibody (orb256458; isotype control, black line) or the rabbit IgG version of J43 (orb758881, blue line) at a dilution of 1:100 for 1h at RT. After washing, the bound antibody was detected using a goat anti-rabbit IgG AlexaFluor® 488 antibody at a dilution of 1:1000 and cells analyzed using a FACSCanto flow-cytometer.

Immunofluorescence staining of mouse splenocytes with anti-PD-1 J43. Immunofluorescence analysis of paraformaldehyde fixed mouse splenocytes on Shi-fix™ coverslips stained with the chimeric rabbit IgG version of J43 (orb758881) at 10 µg/ml for 1h followed by Alexa Fluor® 488 secondary antibody (2 µg/ml), showing membrane staining. The nuclear stain is DAPI (blue). Panels show from left-right, top-bottom orb758881, DAPI, merged channels and an isotype control. The isotype control was an unknown specificity antibody (orb256458) followed by staining with Alexa Fluor® 488 secondary antibody.

Western Blot using Anti-PD-1 antibody J43. Mouse lymph node tissue lysate (35 µg protein in RIPA buffer) were resolved on a SDS PAGE gel and blots were probed with the chimeric rabbit version of J43 (orb758881) at 0.003 µg/ml before detection using an anti-rabbit secondary antibody. A primary incubation of 1h was used and protein was detected by chemiluminescence.