Recombinant Anti-KIFC1 antibody [11445] (ab172620)

Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: KIFC1 knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: HepG2 whole cell lysate (20 µg)

Lanes 1 - 4: Merged signal (red and green). Green - ab172620 (purified) observed at 74 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab172620 was shown to specifically react with KIFC1 in wild-type cells as signal was lost in KIFC1 knockout cells. Wild-type and KIFC1 knockout samples were subjected to SDS-PAGE. Ab172620 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 0.228 µg/ml and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling KIFC1 with purified ab172620 at 1/50 dilution (4.0 µg/ml). Cells were fixed in 100% Methanol. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at 1/200 (2.5 µg/ml) dilution. Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

ab172620 (purified) at 1/20 dilution (1ug) immunoprecipitating KIFC1 in Jurkat whole cell lysates.
Lane 1: Jurkat (Human T cell leukemia T lymphocyte) whole cell lysates 10ug
Lane 2 (+): ab172620 & Jurkat whole cell lysates
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab172620 in Jurkat whole cell lysates
For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.
Blocking and diluting buffer: 5% NFDM/TBST.

Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling KIFC1 with purified ab172620 at 1/20 dilution (10 µg/ml) (red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

Western blot analysis on immunoprecipitation pellet from 293T cell lysate, labeling KIFC1 with ab172620 (unpurified) at 1/10 dilution.

Immunohistochemical analysis of paraffin-embedded Human tonsil tissue laneling KIFC1 with ab172620 (unpurified) at 1/50 dilution.

Immunohistochemical analysis of paraffin-embedded Human cervical carcinoma tissue labeling KIFC1 with ab172620 (unpurified) at 1/50 dilution.

Immunofluorescent staining of HeLa cells labeling KIFC1 with ab172620 (unpurified) at 1/50 dilution.