Lane 1: Wild-type HeLa cell lysate (20µg)
Lane 2: HMGB1 knockout HeLa cell lysate (20µg)
Lanes 1- 2: Merged signal (red and green). Green - ab92310 observed at 30 kDa. Red - loading control ab8245 observed at 37 kDa.
ab92310 Recombinant Anti-HMGB1 antibody [EPR3506] was shown to specifically react with HMGB1 in wild-type HeLa cells in western blot. Loss of signal at the expected size was observed when knockout cell line ab255395 (knockout cell lysate ab263782) was used. The band observed in lane 2 below 25kDa may represent truncated forms and cleaved fragments. Wild-type and HMGB1 knockout samples were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab92310 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4^°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

Lane 1: Wild-type HeLa cell lysate (20µg)
Lane 2: HMGB1 knockout HeLa cell lysate (20µg)
Lanes 1- 2: Merged signal (red and green). Green - ab79823 observed at 30 kDa. Red - loading control ab8245 observed at 37 kDa.
ab79823 Recombinant Anti-HMGB1 antibody [EPR3507] was shown to specifically react with HMGB1 in wild-type HeLa cells in western blot. Loss of signal at the expected size was observed when knockout cell line ab255395 (knockout cell lysate ab263782) was used. The band observed in lane 2 below 25kDa may represent truncated forms and cleaved fragments. Wild-type and HMGB1 knockout samples were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab79823 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4^°C at 1 in 10000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.