Proteasome glo reagent

Manufactured by Promega

Sourced in United States

Proteasome-Glo™ Reagent is a luminescent reagent designed to measure proteasome activity in cellular samples. It contains a substrate that is cleaved by the proteasome, resulting in a luminescent signal proportional to the proteasome activity in the sample.

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Lab products found in correlation

Spectramax i3x microplate reader, by Molecular Devices (1 mentions) Trypsin like, by Promega (1 mentions) Caspase like cell based assays, by Promega (1 mentions) Proteasome glo chymotrypsin like, by Promega (1 mentions) Cyto id autophagy detection kit, by Enzo Life Sciences (1 mentions) Rpmi 1640 medium, by Lonza (1 mentions) Gentamycin, by Lonza (1 mentions)

Close spelling variants of this product

Proteasome-Glo Cell-Based Reagent Proteasome-Glo™ Chymotrypsin-Like Cell-Based Assay Reagent Proteasome-Glo™ Cell-Based Assay Reagent Proteasome-Glo

4 protocols using proteasome glo reagent

Proteasome Activity Assay in NIH3T3 Cells

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NIH3T3(RHO^P23H/GFP), NIH3T3(RHO^WT/GFP) and NIH3T3 cells were seeded in a white-wall clear-bottom 384-well plate at 2500 cells/well in 20 μL of complete mediu. After 3 h of incubation, five μL/well of complete medium containing MTX was added to cells to treat cells with MTX at final concentrations from 0.0195 to 10 μM for 24 h. The cells treated with a complete medium containing 0.1% DMSO were used as the 100% control, and those treated with 5 μM MG-132 for 8 h were used as the 0% control. For endpoint proteasome activity measurement, each well was added with 25 μL of Proteasome-Glo™ Reagent containing the Suc-LLVY-Glo™ substrate (Promega, G8660, Madison, WI USA). The 384-well plate was shaken for 2 min to mix the solutions followed by a 24-min incubation at room temperature. Luminescence of each well was detected by a SpectraMax I3X microplate reader (Molecular Devices). The chymotrypsin-like proteasome activity was normalized by the 100% and 0% controls, respectively.

Liu X., Feng B., Vats A., Tang H., Seibel W., Swaroop M., Tawa G., Zheng W., Byrne L., Schurdak M, & Chen Y. (2020). Pharmacological clearance of misfolded rhodopsin for the treatment of RHO-associated retinitis pigmentosa.. FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 34(8), 10146-10167.

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Proteasome Activity Assay in Cell Extracts

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Cell extracts (25 µg) were incubated with 50 µL of Proteasome-Glo™ Reagent containing Z-LRR-Glo™ Substrate coupled to luciferin (Proteasome-Glo™Tripsin-like assay, Cat No: G8631, Promega Corporation, Madison, WI, USA) for 30 min in dark. Proteasome-induced substrate cleavage generates a “glow-type” luminescent signal produced by the luciferase reaction. Record luminescence signal with a plate-reading luminometer. As a control, 0.25 mM of proteasome inhibitor lactacystin was applied for 5 h prior to the beginning of proteasome activity assays.

Liu X., Balaraman K., Lynch C.C., Hebron M., Shah P.K., Hu S., Stevenson M., Wolf C, & Moussa C. (2022). Inhibition of Ubiquitin-Specific Protease-13 Improves Behavioral Performance in Alpha-Synuclein Expressing Mice. International Journal of Molecular Sciences, 23(15), 8131.

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Proteasomal Activity Profiling in Cells

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The cell-based proteasome activity assay to determine the sub-catalytic proteasomal activities in cells was performed as previously described[45 (link)]. Cells were seeded in solid white 96-well plates 24h prior to treatment. Cells were then treated with the indicated drug (0–50 nM) for 24 h. Cells were incubated for 15 min the proteasome Glo™ reagents according to the manufacturer’s instructions (Promega), Proteasome inhibition of the caspase, trypsin and chymotrypsin activity sites were measured by addition of luminogenic substrates Z-nLPnLD-aminoluciferin, Z-LRR-aminoluciferin and Suc-LLVY-aminoluciferin, respectively.

Pierce M.R., Robinson R.M., Ibarra-Rivera T.R., Pirrung M.C., Dolloff N.G, & Bachmann A.S. (2019). Syrbactin proteasome inhibitor TIR-199 overcomes bortezomib chemoresistance and inhibits multiple myeloma tumor growth in vivo. Leukemia research, 88, 106271.

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Proteasome Activity Assay Protocol

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RPMI-1640 medium and gentamycin were purchased from Lonza. Heat-inactivated low endotoxin fetal bovine serum (FBS) was from Omega scientific. Highly purified, deep rough chemotype LPS (Re-LPS) from E. coli D31m4 was prepared as described previously [14 ]. Proteasome-Glo reagents for determining the cellular proteasome’s activities were purchased from Promega (Madison, WI, USA). Ac-WLA-AMC, Ac-ANW-AMC, and Ac-PAL-AMC fluorogenic substrates, as well as human proteasomes and immunoproteasomes were purchased from R&D systems. Trans-Resveratrol (later called resveratrol) was purchased from “Mega Resveratrol” (60 Newtown Toad # 32, Danbury CT, USA). ONX-0914, specific inhibitor of LMP7 was purchased from UBPBio. MTT (Thiazolyl Blue Tetrazolium Bromide) for cell death assay and PMA (phorbol 12-myristate 13-acetate) for differentiation were purchased from Sigma-Aldrich. Cyto-ID® Autophagy Detection Kit was purchased from Enzo Life Sciences Inc. (Farmingdale, NY). Proteasome-Glo™ Chymo Trypsin-Like, Trypsin-Like and Caspase-Like Cell-Based Assays were purchased from Promega (Madison, WI).

Silswal N., Reddy N.S., Qureshi A.A, & Qureshi N. (2018). RESVERATROL DOWNREGULATES BIOMARKERS OF SEPSIS VIA INHIBITION OF PROTEASOME’S PROTEASES. Shock (Augusta, Ga.), 50(5), 579-588.

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