产品资料

Abcam/Anti-HMGB1 antibody (ab18256)/1/ab18256

Western blot - Anti-HMGB1 antibody (ab18256)
All lanes : Anti-HMGB1 antibody (ab18256) at 1 µg/mlLane 1 : Wild-type HAP1 whole cell lysateLane 2 : HMGB1 knockout HAP1 whole cell lysateLysates/proteins at 20 µg per lane.Predicted band size: 25 kDa
Lanes 1 - 4: Merged signal (red and green). Green - ab18256 observed at 29 kDa. Red - loading control, ab9484, observed at 37 kDa.
ab18256 was shown to recognize HMGB1 in wild-type HAP1 cells as signal was lost at the expected MW in HMGB1 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and HMGB1 knockout samples were subjected to SDS-PAGE. Ab18256 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1 μg/ml and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
Immunocytochemistry/ Immunofluorescence - Anti-HMGB1 antibody (ab18256)
ab18256 stained inHeLa cells. Cells were fixed with 4% paraformaldehyde (10min) at room temperature and incubated with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% triton for 1h at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab18256 at 1µg/ml and ab7291 (Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control) at 1/1000 dilutionovernight at +4°C. The secondary antibodies wereGoat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120)(pseudo-colored red)andGoat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) secondary antibody(colored green)used at 1 ug/ml for 1hour at room temperature. DAPI was used to stain the cell nuclei (colored blue) at a concentration of 1.43µM for 1hour at room temperature
Western blot - Anti-HMGB1 antibody (ab18256)
All lanes : Anti-HMGB1 antibody (ab18256) at 1 µg/mlLane 1 : NIH 3T3 whole cell lysate (ab7179)Lane 2 : MEF1 (Mouse embryonic fibroblast cell line) Whole Cell LysateLane 3 : PC-12 (Rat adrenal pheochromocytoma cell line) Whole Cell LysateLysates/proteins at 10 µg per lane.SecondaryLanes 1 & 3 : IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilutionLane 2 : IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L)at 1/10000 dilutionPerformed under reducing conditions.Predicted band size: 25 kDaObserved band size: 29 kDa
why is the actual band size different from the predicted?Additional bands at: 59 kDa. We are unsure as to the identity of these extra bands.
Western blot - Anti-HMGB1 antibody (ab18256)
All lanes : Anti-HMGB1 antibody (ab18256) at 1 µg/mlLane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysateLane 2 : Jurkat whole cell lysate (ab7899)Lane 3 : A431 whole cell lysate (ab7909)Lane 4 : HEK293 whole cell lysate (ab7902)Lysates/proteins at 10 µg per lane.SecondaryAll lanes : IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilutionPerformed under reducing conditions.Predicted band size: 25 kDaObserved band size: 29 kDa why is the actual band size different from the predicted?
Western blot - Anti-HMGB1 antibody (ab18256)
Anti-HMGB1 antibody (ab18256) at 1 µg/ml + Recombinant Human HMGB1 protein (ab56525) at 0.01 µgSecondaryGoat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilutionDeveloped using the ECL technique.Performed under reducing conditions.Predicted band size: 25 kDaExposure time: 2 minutes
Western blot - Anti-HMGB1 antibody (ab18256)This image is courtesy of an Abreview submitted by Dr Francesca Cerbai
All lanes : Anti-HMGB1 antibody (ab18256) at 1/1000 dilutionLane 1 : Rat brain whole tissue lysate - infused with asf for 1 weekLane 2 : Rat brain whole tissue lysate - infused with LPS for 1 weekLane 3 : Rat brain whole tissue lysate - infused with acsf for 8 weeksLane 4 : Rat brain whole tissue lysate - infused with LPS for 8 weeksLane 5 : Rat brain whole tissue lysate - infused with LPS for 4 weeksLane 6 : Rat brain whole tissue lysate -infused with LPS for 4 weeks, after 2 weeks of LPS infusion were treated with neramexane for the next 2 weeksLane 7 : Rat brain whole tissue lysate - infused with LPS for 4 weeks, after 2 weeks of LPS infusion were treated with memantine for the next 2weeks.Lysates/proteins at 40 µg per lane.SecondaryAll lanes : Biotinylated Goat anti-rabbit IgGDeveloped using the ECL technique.Performed under reducing conditions.Predicted band size: 25 kDaExposure time: 30 seconds
See Abreview

对照品标准品，生物试剂盒，gibco胎牛血清，试剂采购网，中国试剂网，细胞因子试剂盒

Abcam/Anti-HMGB1 antibody (ab18256)/1/ab18256

Images