重组Anti-Ferritin抗体[EPR3004Y] - BSA and Azide free (ab271854)

Immunofluorescence staining of 4% paraformaldehyde fixed, 0.1% Triton X-100 permeabilized Jurkat cells (human T cell leukemia T lymphocyte from peripheral blood) with ab75973 at a working dilution of 1/100. The secondary antibody, ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed, was used at a dilution of 1/1000. The counterstain, ab195889 anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) was used at a dilution of 1/200.

Confocal image showing positive staining in Jurkat cells (shown in green). The counterstain was observed in magenta. Nuclear DNA was labeled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

This data was developed using the same antibody clone in a different buffer formulation (ab75973).

All lanes : Anti-Ferritin antibody [EPR3004Y] (ab75973) at 1/1000 dilution

Lane 1 : Wild-type MCF7 cell lysate
Lane 2 : FTH1 knockout MCF7 cell lysate
Lane 3 : Wild-type HEK-293T ab255553 cell lysate
Lane 4 : FTH1 knockout HEK-293T ab260185 cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 21 kDa
Observed band size: 24 kDa why is the actual band size different from the predicted?

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab75973).

False colour image of Western blot: Anti-Ferritin antibody [EPR3004Y] 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, ab75973 was shown to bind specifically to Ferritin. A band was observed at 24 kDa in wild-type MCF7 cell lysates with no signal observed at this size in FTH1 knockout cell line ab269493 (knockout cell lysate ab269655). To generate this image, wild-type and FTH1 knockout MCF7 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.