Western blot analysis of GAPDH was performed by loading various cell lysates onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a low fluorescence PVDF membrane and blocked with Fish Serum blocking buffer for at least 1 hour. The membrane was probed with a GAPDH Loading Control Monoclonal Antibody (GA1R), Alexa Fluor 488 (Cat. No. MA5-15738-A488) at a dilution of 1:500 for overnight at 4°C on a rocking platform and washed in TBS-0.1% Tween-20. Detection was performed using a fluorescence imaging system.
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is well known for its role in glycolysis. GAPDH catalyzes the phosphorylation of glyceraldehyde-3-phosphate. In addition to this metabolic function, GAPDH is involved in many nonmetabolic cellular processes, including membrane fusion, transcription, RNA transport, DNA replication, and apoptosis. GAPDH has been associated with various cancers and may also play a part in neurodegenerative diseases like Alzheimer’s and Huntington’s diseases. While it is expressed differentially from tissue to tissue, GAPDH is often stably and constitutively expressed, making it useful as a loading control. Quality Invitrogen GAPDH antibodies are available for a variety of research needs. We also offer fluorescent and enzyme-conjugated versions for your added convenience.
Applications
Immunofluorescence analysis of GAPDH (green) showing staining in the cytoplasm of HeLa cells (right), compared with a negative control without primary antibody (left). Cells were probed with a GAPDH Loading Control Monoclonal Antibody (GA1R) (Cat. No. MA5-15738). Cells were washed with PBST and incubated with a secondary antibody conjugated to DyLight 488. F-actin was stained with phalloidin (red), and nuclei were stained with Hoechst stain (blue).
Immunohistochemical analysis of GAPDH showing staining in the nucleus and cytoplasm of paraffin-embedded human kidney tissue (right), compared with a negative control without primary antibody (left). The cells were probed with a GAPDH Loading Control Monoclonal Antibody (GA1R0 (Cat. No. MA5-15738).
Resources
For Research Use Only. Not for use in diagnostic procedures.
For Research Use Only. Not for use in diagnostic procedures.