Anti-CrkL antibody [Y243] - BSA and Azide free

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-CrkL antibody [Y243] - BSA and Azide free (AB247234)

This data was developed using the same antibody clone in a different buffer formulation (ab32126).
Immunocytochemistry analysis of K-562 (human chronic myelogenous leukemia lymphoblast) labeling CrkL with purified ab32126 at 1/100 dilution (10 µg/ml). Cells were fixed with 100% methanol. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/1000 (2 µg/ml) was used as the secondary antibody. ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1/200 (2.6 µg/ml) was used as counterstain. Nuclei were stained blue with DAPI.
Negative control : PBS instead of the primary antibody.

• WB

Lab

Western blot - Anti-CrkL antibody [Y243] - BSA and Azide free (AB247234)

This data was developed using the same antibody clone in a different buffer formulation (ab32126).

Lanes 1-3 : Merged signal (red and green). Green - ab32126 observed at 37 kDa. Red - loading control ab7291 observed at 50 kDa.

ab32126 Anti-CrkL antibody [Y243] was shown to specifically react with CrkL in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265993 (knockout cell lysate ab257397) was used. Wild-type and CrkL knockout samples were subjected to SDS-PAGE. ab32126 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-CrkL antibody [Y243] (<a href='/en-us/products/primary-antibodies/crkl-antibody-y243-ab32126'>ab32126</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

CRKL knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human CRKL knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-crkl-knockout-hela-cell-line-ab265993'>ab265993</a>)

Lane 3:

K-562 cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 34 kDa

Observed band size: 37 kDa

false