Anti-TCF12 抗体 (ab245540)

All lanes : Anti-TCF12 antibody (ab245540) at 1/1000 dilution

Lane 1 : Wild-type Jurkat cell lysate
Lane 2 : TCF12 CRISPR-Cas9 edited Jurkat cell lysate
Lane 3 : HeLa cell lysate
Lane 4 : MOLT-4 cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 73 kDa
Observed band size: 85 kDa why is the actual band size different from the predicted?

Western blot: Anti-TCF12 antibody staining at 1/1000 dilution, shown in black; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab245540 was shown to bind specifically to TCF12. A band was observed at 85 kDa in wild-type Jurkat cell lysates with no signal observed at this size in TCF12 CRISPR-Cas9 edited cell line ab274931 (CRISPR-Cas9 edited cell lysate ab274989). The band observed in the CRISPR-Cas9 edited lysate lane below 85 kDa is likely to represent a truncated form of TCF12. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and TCF12 CRISPR-Cas9 edited Jurkat cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% milk in TBS-0.1% Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times before development with Optiblot (ECL reagent ab133456) and imaged with 20 minutes exposure time. Secondary antibodies used were HRP conjugated Goat anti-Rabbit (H+L) and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes : Anti-TCF12 antibody (ab245540) at 0.1 µg/ml

Lane 1 : HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 50 µg
Lane 2 : HeLa whole cell lysate at 15 µg
Lane 3 : HeLa whole cell lysate at 5 µg

Developed using the ECL technique.

Predicted band size: 73 kDa


Exposure time: 3 minutes

TCF12 was immunoprecipitated from HeLa (human epithelial cell line from cervix adenocarcinoma) whole cell lysate (1 mg for IP, 20% of IP loaded) with ab245540 at 3 µg/mg lysate. Western blot was performed from the immunoprecipitate using ab245540 at 1 µg/ml.

Lane 1: ab245540 IP in HeLa whole cell lysate.
Lane 2: Control IgG IP in HeLa whole cell lysate.

Detection: Chemiluminescence with exposure time of 10 seconds.