Recombinant Anti-Dnmt3a antibody [EPR18455] - BSA and Azide free (ab232391)

This data was developed using the same antibody clone in a different buffer formulation (ab188470).

Lanes 1-4: Merged signal (red and green). Green - ab188470 observed at 125 kDa. Red - loading control ab8245 observed at 37 kDa.

 ab188470 Anti-Dnmt3a antibody [EPR18455] was shown to specifically react with Dnmt3a in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab261793 (knockout cell lysate ab257128) was used. Wild-type and Dnmt3a knockout samples were subjected to SDS-PAGE. ab188470 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 2000 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.

Lanes 1 - 4: Merged signal (red and green). Green - ab188470 observed at 125 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab188470 was shown to recognize Dnmt3a when Dnmt3a knockout samples were used, along with additional cross-reactive bands. Wild-type and Dnmt3a knockout samples were subjected to SDS-PAGE. ab188470 and ab8245 (loading control to GAPDH) were diluted to 1/5000 and 1/10000 respectively and incubated overnight at 4°C. 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 1/10000 dilution for 1 hour at room temperature before imaging.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab188470).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human placenta tissue sections labeling Dnmt3a with purified ab188470 at 1/2000 (0.409 µg/ml). Antigen retrieval was heat mediated using ab93684 (Tris/EDTA buffer, pH 9.0). Goat Anti-Rabbit IgG H&L (HRP) was used as the secondary antibody. Hematoxylin was used as a counterstain.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab188470).

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cells from cervix adenocarcinoma) cells labeling Dnmt3a with ab188470 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing nuclear and weakly cytoplasmic staining on HeLa cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

The negative controls are as follows:
-ve control 1: ab188470 at 1/1000 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab188470).

Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling Dnmt3a with purified ab188470 at 1/80 dilution (10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control. 

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab188470). 

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat spleen tissue sections labeling Dnmt3a with purified ab188470 at 1/2000 (0.409 µg/ml). Antigen retrieval was heat mediated using ab93684 (Tris/EDTA buffer, pH 9.0). Goat Anti-Rabbit IgG H&L (HRP) was used as the secondary antibody. Hematoxylin was used as a counterstain.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab188470).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse testis tissue sections labeling Dnmt3a with purified ab188470 at 1/2000 (0.409 µg/ml). Antigen retrieval was heat mediated using ab93684 (Tris/EDTA buffer, pH 9.0). Goat Anti-Rabbit IgG H&L (HRP) was used as the secondary antibody. Hematoxylin was used as a counterstain.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab188470).