Immunofluorescent analysis of Proteasome 20S LMP7 (green) in HeLa cells. The cells were fixed with 4% paraformaldehyde for 15 minutes, permeabilized with 0.1% Triton X-100 in TBS for 10 minutes, and blocked with 3% Blocker BSA (Product # 37525) in PBS for 15 minutes at room temperature. Cells were stained with or without Proteasome 20S LMP7 rabbit polyclonal antibody (Product # PA1-972), at a concentration of 5ug/ml for 1 hour at room temperature, and then incubated with a Alexa Fluor® 488 Superclonal goat anti-rabbit IgG secondary antibody (Product # A27034) at a dilution of 1:1000 for 1 hour s at room temperature (both panels, green). Nuclei (both panels, blue) were stained with Hoechst 33342 dye (Product # 62249). Images were taken on a Thermo Scientific ToxInsight at 20X magnification.
|Tested species reactivity||Human|
|Published species reactivity||Human|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic Peptide: C V(259) E S T D V S D L L H Q Y R E A(274)|
|Purification||Antigen affinity chromatography|
|Storage buffer||PBS with 1mg/ml BSA|
|Contains||0.05% sodium azide|
|Storage Conditions||-20° C, Avoid Freeze/Thaw Cycles|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||3-5 µg/10^6 cells|
|Western Blot (WB)||1-3 µg/ml|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
|Western Blot (WB)||See 2 publications below|
PA1-972 detects Proteasome 20S LMP7 in western blot and immunofluorescence.
Proteolytic degradation is critical to the maintenance of appropriate levels of short-lived and regulatory proteins as important and diverse as those involved in cellular metabolism, heat shock and stress response, antigen presentation, modulation of cell surface receptors and ion channels, cell cycle regulation, transcription, and signalling factors. The ubiquitin-proteasome pathway deconstructs most proteins in the eukaryotic cell cytosol and nucleus. Others are degraded via the vacuolar pathway which includes endosomes, lysosomes, and the endoplasmic reticulum.
The 26S proteasome is an ATP-dependent, multisubunit (~31), barrel-shaped molecular machine with an apparent molecular weight of ~2.5 MDa. It consists of a 20S proteolytic core complex which is crowned at one or both ends by 19S regulatory subunit complexes. The 19S regulatory subunits recognize ubiquitinated proteins and play an essential role in unfolding and translocating targets into the lumen of the 20S subunit. An enzymatic cascade is responsible for the attachment of multiple ubiquitin molecules to lysine residues of proteins targeted for degradation. Several genetic diseases are associated with defects in the ubiquitin-proteasome pathway. Some examples of affected proteins include those linked to cystic fibrosis, Angelman and quote;s syndrome, and Liddle syndrome.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Glucosamine induces cell death via proteasome inhibition in human ALVA41 prostate cancer cell.
PA1-972 was used in western blot to investigate the mechanism for glucosamine-induced cell apoptosis in cancer cells
|Liu BQ,Meng X,Li C,Gao YY,Li N,Niu XF,Guan Y,Wang HQ||Experimental and molecular medicine (43:487)||2011|
Inhibition of apoptosis in acute promyelocytic leukemia cells leads to increases in levels of oxidized protein and LMP2 immunoproteasome.
PA1-972 was used in western blot to study the mechanism for the observed accumulation of oxidized proteins during aging and in age-related diseases.
|Khan MA,Oubrahim H,Stadtman ER||Proceedings of the National Academy of Sciences of the United States of America (101:11560)||2004|