SARS-CoV-2 PLpro at a final concentration of 50 nM was added to recombinant human pro-ISG15 (final concentration 350 nM) solution in the presence or absence of biflavones. The reaction buffer was 20 mM Tris (pH 8.0) containing 10 mM DTT and 150 mM NaCl. After a 10 min incubation at 37 °C, the deISGylation reaction was terminated by the addition of 4X LDS loading buffer, followed by heating at 70 °C for 10 min and finally resolving by SDS-PAGE. The protein bands were stained with eLuminol protein gel stain (Genecopoeia, MD, USA), visualized under a UV transilluminator and quantified by ImageLab 3.0 software (Bio– Rad, CA, USA).
Eluminol protein gel stain
Manufactured by GeneCopoeia
Sourced in United States
ELuminol is a protein gel stain designed for the detection and visualization of proteins in polyacrylamide gels. It is a fluorescent dye that binds to proteins, allowing them to be visualized under ultraviolet (UV) or blue light illumination.
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Lab products found in correlation
2 protocols using eluminol protein gel stain
1
SARS-CoV-2 PLpro Deubiquitination Assay
2
Recombinant GUS Production and Purification
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Recombinant human GUS was produced from a CHO-K1 cell line as reported previously (16) . Unpurified GUS from the spent media was either dialyzed against buffer (10 mM Tris-HCl, pH 8.0, with 10% glycerol) or purified as described previously (17) . Cathepsin D (21940130U; Fisher Scientific, Pittsburgh, PA), peptide-N-glycosidase F (PNGase F) (P0704; New England Biolabs, Ipswich, MA) and alkaline phosphatase (10108146001; Sigma Aldrich, St. Louis, MO) were obtained from the respective manufacturers. For SDS-PAGE, Criterion TGX precast gels (5671094 or 5671095; Bio-Rad, Hercules, CA) were used. The following specific stains were purchased: Phos-Tag Phosphoprotein Gel Stain (P005A and P005B; GeneCopoeia, Rockville, MD), eLuminol Protein Gel Stain (P003A, GeneCopoeia), and PageBlue Protein Staining Solution (24620; Thermo Scientific, Waltham, MA). All buffers and aqueous reagents were prepared using deionized and filtered Milli-Q water (specific resistance: 18.2 MΩ/cm).
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