Recombinant Anti-Mucin 5AC antibody [45M1] (ab3649)

Fluorescence multiplex immunohistochemical analysis of the human stomach (Formalin/PFA-fixed paraffin-embedded sections). Panel A: merged staining of anti-Mucin 5AC (ab3649, gray; Opal™690), anti-MUC-6 (ab223846, green; Opal™520) and anti-Rab3D (ab128997, red; Opal™570) on human stomach. Panel B: anti-MUC-6 stained on mucous neck cells. Panel C: anti-Mucin 5AC stained on surface mucous cells. Panel D: anti-Rab3D stained on Chief cells. Opal Polymer HRP Ms + Rb was used as a secondary antibody. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument with an Opal™ 4-color kit. The section was incubated in three rounds of staining: in the order of ab3649 (1/5000 dilution), ab223846 (1/1000 dilution), and ab128997 (1/10000 dilution) for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) was used for 20 mins. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Leica SP8 confocal microscope.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human stomach tissue labeling Mucin 5AC with ab3649 at 1/5000 dilution. Heat mediated antigen retrieval was performed using Citrate buffer pH 6 (epitope retrieval solution 1) for 20 minutes. The section was incubated with ab3649 for 30 mins at room temperature. Goat Anti-Mouse IgG H&L (HRP polymer) (ab214879) was used as the secondary antibody. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Immunocytochemistry analysis of A549 (human lung carcinoma cell line) cells labeling Mucin 5AC with ab3649 at 1/100 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) (ab150113) at 1/1000 was used as the secondary antibody (green). Cells were counterstained with Anti-beta Tubulin rabbit monoclonal antibody (ab179513) at 1/200 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) (ab150080) at 1/1000 dilution (red). Nuclear DNA was labelled with DAPI (blue).

Negative control 1: ab3649 (Mouse monoclonal antibody to Mucin 5AC at 1/100 dilution) and ab150080 (anti-Rabbit secondary Alexa Fluor^® 594 at 1/1000 dilution)

Negative control 2: ab179513 (Rabbit monoclonal antibody to beta Tubulin at 1/200 dilution) and ab150113 (anti-Mouse secondary Alexa Fluor^® 488 at 1/1000 dilution)

Immunohistochemistry (Frozen sections) analysis of mouse stomach tissue labeling Mucin 5AC with ab3649 at 1/100 dilution. Tissue was fixed with 4% paraformaldehyde and permeabilized with 0.2% Triton X-100. Heat mediated antigen retrieval was performed using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20). Goat Anti-Mouse IgG H&L (Alexa Fluor^® 488) (ab150113) was used as the secondary antibody at 1/1000 dilution (green). Nuclei counterstained with DAPI (blue).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse stomach tissue labeling Mucin 5AC with ab3649 at 1/5000 dilution. Heat mediated antigen retrieval was performed using Citrate buffer pH 6 (epitope retrieval solution 1) for 20 minutes. The section was incubated with ab3649 for 30 mins at room temperature. Goat Anti-Mouse IgG H&L (HRP polymer) (ab214879) was used as the secondary antibody. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Immunohistochemistry (Frozen sections) analysis of rat stomach tissue labeling Mucin 5AC with ab3649 at 1/100 dilution. Tissue was fixed with 4% paraformaldehyde and permeabilized with 0.2% Triton X-100. Heat mediated antigen retrieval was performed using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20). Goat Anti-Mouse IgG H&L (Alexa Fluor^® 488) (ab150113) was used as the secondary antibody at 1/1000 dilution (green). Nuclei counterstained with DAPI (blue).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat stomach tissue labeling Mucin 5AC with ab3649 at 1/5000 dilution. Heat mediated antigen retrieval was performed using Citrate buffer pH 6 (epitope retrieval solution 1) for 20 minutes. The section was incubated with ab3649 for 30 mins at room temperature. Goat Anti-Mouse IgG H&L (HRP polymer) (ab214879) was used as the secondary antibody. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument

This image was generated from the hybridoma version of the product.

Immunohistochemical analysis of rat stomach tissue, staining Mucin 5AC with ab3649.

Antigen retrieval was performed via heat mediation in a citrate buffer. Samples were blocked before incubating with primary antibody (1/100) for 1 hour at room temperature. An HRP-conjugated anti-mouse IgG was used as the secondary antibody and staining was detected using DAB.