Key features and details
- Mouse monoclonal [AC-15] to beta Actin
- Suitable for: ICC/IF, WB
- Knockout validated
- Reacts with: Mouse, Rat, Cow, Dog, Human, African green monkey, Chinese hamster
- Isotype: IgG1
Product nameAnti-beta Actin antibody [AC-15]
See all beta Actin primary antibodies
DescriptionMouse monoclonal [AC-15] to beta Actin
Tested Applications & Species
See all applications and species data
Application Species ICC/IFHuman WBHuman
EpitopeN-terminal of the beta isoform of actin.
- ICC/IF: SV40LT-SMC cells WB: HAP1, HeLa, Jurkat, A431, HEK-293, COS-7, NIH/3T3, PC-12, Rat2, CHO, MDBK and MDCK cell lysates. IHC-Fr: Human stomach tissue. ICC/IF: SV40LT-SMC cells.
Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
Storage bufferpH: 7.40
Preservative: 0.097% Sodium azide
Concentration information loading...
Purification notesPurified from hybridoma cell culture.
ab6276 staining beta Actin in SV40LT-SMC cells. The cells were fixed with 4% formaldehyde (10 min), permeabilized in 0.1% Triton X-100 for 5 minutes and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab6276 at a working concentration of 5μg/ml and ab190573, Rabbit monoclonal [EP1332Y] to alpha Tubulin (Alexa Fluor® 647, shown in red) at 1/250 overnight at +4°C, followed by a further incubation at room temperature for 1h with an anti-mouse AlexaFluor® 488 (ab150117) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
Lane 1: Wild-type HAP1 cell lysate (20 µg)
Lane 2: Beta actin knockout HAP1 cell lysate (20 µg)
Lanes 1 and 2: Merged signal (red and green). Green - beta actin, ab6276 observed at 42 kDa. Red - loading control, ab181602 observed at 37 kDa.
Ab6276 was shown to specifically react with beta actin in wild-type HAP1 cells. No band was observed when beta actin knockout samples were used. Wild-type and beta actin knockout samples were subjected to SDS-PAGE. ab6276 (beta actin) and ab181602 (loading control to GAPDH) were diluted 1/5000 and 1/10 000 and incubated overnight at 4°C. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1/10 000 dilution for 1 h at room temperature before imaging.
Western Blot of ab6276 (used as loading control) with whole tissue lysate of grey matter from BA20 (temporal cortex). Ab6276 was diluted 1/50000 and incubated with the sample for 16 hours at 4°C. 5 µg of lysate was loaded onto the gel, which was blocked with 5% milk for 1 hour at 20°C. An Alexa Fluor® 680 conjugated goat anti-mouse antibody, diluted 1/5000, was used as the secondary.
Bands below actin in image are synaptophysin (SYN).
All lanes : Anti-beta Actin antibody [AC-15] (ab6276) at 1 µg/ml
Lane 1 : HeLa (human epithelial cell line from cervix adenocarcinoma) cell lysate
Lane 2 : Jurkat (human T cell leukemia cell line from peripheral blood) cell lysate
Lane 3 : COS-7 (african green monkey kidney fibroblast-like cell line) cell lysate
Lane 4 : NIH/3T3 (mouse embryonic fibroblast cell line) cell lysate
Lane 5 : PC-12 (rat adrenal gland pheochromocytoma cell line) cell lysate
Lane 6 : Rat2 (rat fibroblast cell line) cell lysate
Lane 7 : CHO (chinese hamster ovary cell line) cell lysate
Lane 8 : MDBK (bovine kidney cell line) cell lysate
Lane 9 : MDCK (canine kidney cell line) cell lysate
All lanes : Goat Anti-Mouse IgG-Peroxidase
Developed using the ECL technique.
Predicted band size: 42 kDa
All lanes : Anti-beta Actin antibody [AC-15] (ab6276) at 1/5000 dilution
Lane 1 : HeLa nuclear
Lane 2 : HeLa whole cell lysate
Lane 3 : A431 cell lysate
Lane 4 : Jurkat cell lysate
Lane 5 : HEK293 cell lysate
Lysates/proteins at 20 µg per lane.
All lanes : Alexa Fluor anti mouse at 1/5000 dilution
Performed under reducing conditions.
Predicted band size: 42 kDa
Observed band size: 42 kDa
MDCK cells induced with increasing amounts of doxycycline to control expression of the gene of interest. All cells were normalized for loading with an albumin protein standard assay. Anti-beta actin (ab6276) was used at a concentration of 1:5000 in a milk blocking solution. B-actin blotting confirms the albumin assay in showing that an equal amount of lysate was loaded in each lane.