Mouse Monoclonal GAPDH Antibody

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     All lanes : Anti-GAPDH Antibody at 1:8000 dilution Lane 1: Hela whole cell lysate Lane 2: Jurkat whole cell lysate Lane 3: A549 whole cell lysate Lysates/proteins at 20 µg per lane. Secondary Goat Anti-mouse IgG,  (H+L), Peroxidase conjugated at 1/10000 dilution. Predicted band size : 36 kDa Blocking/Dilution buffer: 5% NFDM/TBST.    

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     Overlay histogram showing Hela cells stained with P33889(green line).  The cells were fixed with 2% paraformaldehyde (10 min) and then permeabilized with 90% methanol for 10 min.  The cells were then icubated in 2% bovine serum albumin to block non-specific protein-protein interactions followed by the antibody (P33889,  1:25 dilution) for 60 min at 37ºC.  The secondary antibody used was Goat-Anti-Mouse IgG, DyLight® 488 Conjugated Highly Cross-Adsorbed(OJ192088) at 1/200 dilution for 40 min at 37ºC.  Isotype control antibody (blue line) was mouse IgG1 (1μg/1x10^6 cells) used under the same conditions.  Acquisition of >10, 000 events was performed.    

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     P33889 staining GAPDH in human kidney tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed,  paraffin-embedded sections).  Tissue was fixed with formaldehyde and blocked with 3% BSA for 0. 5 hour at room temperature; antigen retrieval was by heat mediation with a citrate buffer (pH6).  Samples were incubated with primary antibody (1/25) for 1 hours at 37°C.  A undiluted biotinylated goat polyvalent antibody was used as the secondary antibody.    

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     P33889 staining GAPDH in human colon tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed,  paraffin-embedded sections).  Tissue was fixed with formaldehyde and blocked with 3% BSA for 0. 5 hour at room temperature; antigen retrieval was by heat mediation with a citrate buffer (pH6).  Samples were incubated with primary antibody (1/25) for 1 hours at 37°C.  A undiluted biotinylated goat polyvalent antibody was used as the secondary antibody.    

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a highly conserved enzyme critical to glycolysis, catalyzing the oxidation and phosphorylation of glyceraldehyde-3-phosphate. Beyond its metabolic role, GAPDH participates in non-glycolytic processes, including DNA repair, RNA transport, and apoptosis. Due to its constitutive expression across tissues and cell types, GAPDH is widely used as a "housekeeping" protein reference in molecular biology.

GAPDH antibodies are essential tools for detecting this protein in techniques like Western blotting, immunofluorescence, and ELISA. They are frequently employed as loading controls to normalize data, ensuring equal protein loading across samples. These antibodies typically target conserved epitopes, enabling cross-reactivity with GAPDH orthologs from various species, including humans, mice, and rats.

Despite its popularity, reliance on GAPDH as a control requires caution. Certain experimental conditions—such as hypoxia, oxidative stress, or disease states—may alter GAPDH expression levels, potentially compromising its reliability. Researchers are advised to validate its stability under specific experimental settings. Commercially available GAPDH antibodies vary in clonality (monoclonal/polyclonal), host species, and conjugate labels, allowing flexibility for diverse applications.

In summary, GAPDH antibodies serve as indispensable reagents for normalizing protein studies, though contextual validation remains crucial to ensure experimental accuracy.