All lanes : Anti-c-Myc antibody (ab39688) at 1/500 dilution

Lane 1 : NIH/3T3 cell extracts
Lane 2 : NIH/3T3 cell extracts with synthesized peptide

Lysates/proteins at 40 µg per lane.

Predicted band size: 49 kDa
Observed band size: 49 kDa

Blocking buffer: 5% (w/v) non-fat dry milk in TBST.

Primary antibody dilution buffer: 5%(w/v)non-fat dried milk,0.1%(v/v), Tween-20 in TBST.

Secondary antibody dilution buffer: 5%(w/v)non-fat dried milk,0.1%(v/v),Tween-20 in TBST.

12% SDS gel. Nitrocellulose membrane. 

Blocking: Room temperature for 2 hours or overnight at 4°C. Then wash 3x for 5 minutes with 0.05% blocking buffer.

Primary antibody incubation: diluted in TBST at 1/500. Incubate overnight with 4 degrees shaking. Then, in 0.05% TBST, wash membrane 3-4 times for 10min.

Secondary antibody incubation: diluted in TBST at 1/2000. Incubate 37°C for 1 hour. Then, in 0.05% TBST, wash membrane 3-4 times for 10min.

ECL development. 

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse pancreatic cancer tissue sections labeling c-Myc with ab39688 at 1/50 dilution. Tissue was fixed in paraformaldehyde and permeabilized with TBST. Heat mediated antigen retrieval was performed using a citrate buffer. Tissue was blocked in 10% serum for 1 hour at 20°C. A polyclonal goat anti-rabbit biotin conjugated secondary antibody was used at 1/1000 dilution.  

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human pancreatic cancer tissue sections labeling c-Myc with ab39688 at 1/50 dilution. Tissue was fixed in paraformaldehyde and permeabilized with TBST. Heat mediated antigen retrieval was performed using a citrate buffer. Tissue was blocked in 10% serum for 1 hour at 20°C. A polyclonal goat anti-rabbit biotin conjugated secondary antibody was used at 1/1000 dilution.  

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ab39688 staining P22 mouse dorsal skin tissue sections by IHC-P.  Sections were PFA fixed and subjected to heat mediated antigen retrieval in citrate buffer prior to blocking with 10% serum for 1 hour at RT.  The primary antibody was diluted 1/20 and incubated with the sample for 24 hrs at 4°C.  A biotinylated goat anti-rabbit antibody was used as the secondary in conjunction with streptavidin-Cy3®.

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ab39688, at 1/50 dilution, staining Human c-Myc in breast carcinoma tissue in the absence (left) and presence (right) of blocking peptide by Immunohistochemistry, Paraffin embedded tissue.