Anti-Ferritin Light Chain antibody [CPT-R10-58-11]

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Ferritin Light Chain antibody [CPT-R10-58-11] (AB243096)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized HepG2 (human liver hepatocellular carcinoma cell line) cell line labeling Ferritin Light Chain with ab243096 at 1/500 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody.

• WB

Lab

Western blot - Anti-Ferritin Light Chain antibody [CPT-R10-58-11] (AB243096)

Western blot : Anti-Ferritin Light Chain antibody [CPT-R10-58-11] ab243096 staining at 1/1000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 19 kDa in Wild-type HeLa cell lysates with no signal observed at this size in FTL knockout HeLa cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5% 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 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Ferritin Light Chain antibody [CPT-R10-58-11] (ab243096) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysates at 20 µg

Lane 2:

Western blot - Human FTL knockout HeLa cell line (<a href='/ko/products/cell-lines/human-ftl-knockout-hela-cell-line-ab265533'>ab265533</a>) at 20 µg

Lane 3:

HepG2 cell lysates at 20 µg

Lane 4:

T-47D cell lysates at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 20 kDa

Observed band size: 19 kDa

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• WB

Unknown

Western blot - Anti-Ferritin Light Chain antibody [CPT-R10-58-11] (AB243096)

Exposure time : Lane 1-4 : 26 seconds, Lane 5 : 5.5 seconds.

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Ferritin Light Chain antibody [CPT-R10-58-11] (ab243096) at 1/1000 dilution

Lane 1:

NCI-H460 (human lung cancer cell line) whole cell lysate at 20 µg

Lane 2:

HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg

Lane 3:

MCF7 (human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Lane 4:

HepG2 (human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg

Lane 5:

Human brain lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/ko/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 20 kDa

Observed band size: 19 kDa

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• ELISA

Unknown

ELISA - Anti-Ferritin Light Chain antibody [CPT-R10-58-11] (AB243096)

50ng CPTAC-14d (1000ng/mL, 50μL per well) was coated onto 96-wells. Serial dilutions (0, 0.2, 0.9, 3.9, 15, 62, 250, 1000ng/mL) of ab243096 (50μL) were incubated with CPTAC-14d in each well for 60 minutes by shaking. This was followed by adding goat anti-rabbit IgG, (H+L), phosphatase-conjugated secondary antibody (50μL) into each well and incubating for another 40 minutes. After washing, 50μL of PNPP was added and incubated for 15 minutes without shaking for color development. OD was read at 405nm within 5 minutes.