Recombinant Anti-Bcl-2 antibody [E17] - BSA and Azide free (ab185002)

This data was developed using the same antibody clone in a different buffer formulation (ab32124).

Lanes 1-4: Merged signal (red and green). Green - ab32124 observed at 26 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab32124 was shown to react with Bcl-2 in wild-type HeLa. Loss of signal was observed when knockout sample ab263752 was used. Wild-type and Bcl-2 knockout samples were subjected to SDS-PAGE. ab32124 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) 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.

Blocking buffer and concentration: 5% NFDM/TBST

Diluting buffer and concentration: 5% NFDM/TBST

Methanol-fixed HeLa (human epithelial cell line from cervix adenocarcinoma) cells stained for Bcl-2 using ab32124 at 1/200 dilution in ICC/IF, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor­ 488) preadsorbed (ab150081). Counter-stained with DAPI.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

Formaldehyde-fixed, paraffin-embedded human DLBCL U2932 cell line xenograft tissue stained for Bcl-2 using ab32124 at 1/200 dilution in immunohistochemical analysis, followed by Goat anti-Rabbit IgG Alexa Fluor^® 555.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

Bcl-2 expression determined by immunohistochemical analyses of the 4 human UM xenografts (between 3 to 5 tumors have been studied per condition).

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

ab32124 (purified) at 1/30 immunoprecipitating Bcl-2 in Jurkat (human T cell leukemia cell line from peripheral blood) cell lysate (Lane 1). Lane 2 - PBS. For western blotting, a HRP-conjugated anti-rabbit IgG, specific to the non-reduced form of IgG was used as the secondary antibody (1/1500).

Blocking buffer and concentration: 5% NFDM/TBST.

Diluting buffer and concentration: 5% NFDM /TBST.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

Immunocytochemistry/Immunofluorescence analysis of MCF7 (human breast adenocarcinoma cell line) cells labelling Bcl-2 with purified ab32124 at 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. The cells were co-stained with ab7291, a mouse anti-tubulin (1/500) using ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/500) as the secondary antibody. Nuclei counterstained with DAPI (blue). 

Control 1: primary antibody (1/100) and secondary antibody, ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/500).

Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/500).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human B-cell lymphoma tissue labelling Bcl-2 with purified ab32124 at 1/250. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

Immunocytochemistry/Immunofluorescence analysis of MCF7 (human breast adenocarcinoma cell line) cells labelling Bcl-2 with unpurified ab32124 at 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. An Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. The cells were co-stained with ab7291, a mouse anti-tubulin antibody (1/500) using ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/500) as the secondary. Nuclei counterstained with DAPI (blue).

Control: primary antibody (1/100) and secondary antibody, ab150120, an Alexa Fluor^® 594-conjugated goat anti-mouse IgG (1/500).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast carcinoma tissue labelling Bcl-2 with unpurified ab32124 at 1/200 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human B cell lymphoma tissue labelling Bcl-2 with unpurified ab32124.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

Immunohistochemical analysis of Human salivary glands taken from patients with primary Sjögren's syndrome, staining Bcl-2 with unpurified ab32124.

Antigen retrieval was performed via heat mediation in a citrate buffer (pH 6). Sections were blocked using 2% BSA, 10% normal serum and permeabilized with 0.5% Triton X-100. Samples were incubated with primary antibody (1/100) for one hour at room temperature. An Alexa Fluor^® 594-conjugated anti-rabbit IgG was used as the secondary antibody.
N.B. Panels B and D are higher magnifications of panels A and C, respectively.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).

Equilibrium disassociation constant (K_D)

Click here to learn more about K_D

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32124).