Anti-Flt3 / CD135 抗体 [EPR22232-318] - BSA and Azide free (ab245824)

All lanes : Anti-Flt3 / CD135 antibody [EPR22232-318] (ab245116) at 1/1000 dilution

Lane 1 : Wild-type THP-1 cell lysate
Lane 2 : FLT3 knockout THP-1 cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 113 kDa
Observed band size: 130 kDa why is the actual band size different from the predicted?

False colour image of Western blot: Anti-Flt3 / CD135 antibody [EPR22232-318] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab245116 was shown to bind specifically to Flt3 / CD135. A band was observed at 130 kDa in wild-type THP-1 cell lysates with no signal observed at this size in FLT3 knockout cell line ab282857 (knockout cell lysate ab283225). The band observed at 60 kDa has not been identified. To generate this image, wild-type and FLT3 knockout THP-1 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 then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab245116).

Flt3 / CD135 was immunoprecipitated from 0.35 mg of THP-1 (human monocytic leukemia cell line) whole cell lysate with ab245116 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab245116 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution.

Lane 1: THP-1 whole cell lysate 10 μg (Input).
Lane 2: ab245116 IP in THP-1 whole cell lysate (+).
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab245116 in THP-1 whole cell lysate (-).

Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 1 minute.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab245116).