Anti-CD74 antibody [CLIP/3127R] - BSA and Azide free

• Flow Cyt

Lab

Flow Cytometry - Anti-CD74 antibody [CLIP/3127R] - BSA and Azide free (AB270296)

This data was developed using the same antibody clone in a different buffer formulation (ab270265).

Flow cytometry overlay histogram showing wild-type Raji (green line) and CD74 knockout Raji cells (ab273378) stained with ab270265 (red line). The cells were incubated in 1x PBS containing 10μg/ml human IgG and 10% normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody (ab270265) (1x10⁶ in 100μl at 1 μg/ml) for 30 min at 4°C.

The secondary antibody Goat anti-rabbit IgG H&L (Alexa Fluor^® 488, pre-adsorbed) (ab150081) was used at 1/2000 for 30 min at 4°C.

Isotype control antibody was Rabbit IgG (monoclonal) (ab172730) used at the same concentration and conditions as the primary antibody (wild-type Raji cells - black line; CD74 knockout Raji cells ab273378 - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

• WB

Lab

Western blot - Anti-CD74 antibody [CLIP/3127R] - BSA and Azide free (AB270296)

This data was developed using the same antibody clone in a different buffer formulation (ab270265).

Lanes 1 - 4 : Merged signal (red and green). Green - ab270265 observed at 35 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

ab270265 was shown to react with CD74 in western blot. The band observed in CD74 CRISPR/Cas9 edited cell line ab273378 (CRISPR/Cas9 edited lysate ab275529) below 35 kDa is likely to represent a truncated form. This has not been investigated further. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween^®) before incubation with ab270265 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at 1 μg/ml and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-CD74 antibody [CLIP/3127R] (<a href='/en-us/products/primary-antibodies/cd74-antibody-clip-3127r-ab270265'>ab270265</a>) at 1 µg/mL

Lane 1:

Wild-type Raji cell lysate at 30 µg

Lane 2:

CD74 knockout Raji cell lysate at 30 µg

Lane 2:

Western blot - Human CD74 knockout Raji cell line (<a href='/en-us/products/cell-lines/human-cd74-knockout-raji-cell-line-ab273378'>ab273378</a>)

Lane 3:

Jurkat cell lysate at 30 µg

Lane 4:

HepG2 cell lysate at 30 µg

Predicted band size: 34 kDa

Observed band size: 35 kDa

false

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD74 antibody [CLIP/3127R] - BSA and Azide free (AB270296)

Formalin-fixed, paraffin-embedded human tonsil tissue stained for CD74 using ab270265 at 2 μg/ml in immunohistochemical analysis.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA and sodium azide (ab270265).

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-CD74 antibody [CLIP/3127R] - BSA and Azide free (AB270296)

Flow cytometric analysis of Raji (Human Burkitt's lymphoma cell line) cells labeling CD74 using ab270265 at 2μg/10⁶ cells followed by goat anti-mouse IgG-CF488 (Blue); isotype control (Red).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA and sodium azide (ab270265).

• WB

Supplier Data

Western blot - Anti-CD74 antibody [CLIP/3127R] - BSA and Azide free (AB270296)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA and sodium azide (ab270265).

All lanes:

Western blot - Anti-CD74 antibody [CLIP/3127R] - BSA and Azide free (ab270296) at 2 µg/mL

All lanes:

HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) cell lysate

Predicted band size: 34 kDa

false