All lanes : Anti-Ferritin Light Chain antibody [FTL/1386] (ab218400) at 1/1000 dilution

Lane 1 : Wild-type HeLa cell lysate
Lane 2 : Ferritin Light Chain knockout HeLa cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 20 kDa
Observed band size: 20 kDa

Lanes 1- 2: Merged signal (red and green). Green - ab218400 observed at 20 kDa. Red - Anti-alpha Tubulin antibody [EP1332Y] - Microtubule Marker (ab52866) observed at 50 kDa.

 ab218400 was shown to react with Ferritin Light Chain in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab265534 (knockout cell lysate ab256927) was used. Wild-type HeLa and Ferritin Light Chain knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab218400 and Anti-alpha Tubulin antibody [EP1332Y] - Microtubule Marker (ab52866) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.

Allele-1: 1 bp insertion in exon 1.

 

Allele-2: 2 bp insertion in exon 1.

 

Representative images of FTL knockout HeLa cells, low and high confluency examples (top left and right respectively) and wild-type HeLa cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using a EVOS XL Core microscope.