Tyrosinase zymography was executed as described previously9 . Cells (1 × 105 cells/mL) were treated with or without NNFE for 3 d. Next, cells were rinsed with PBS twice and collect the cell lysate using RIPA buffer affixed with protease and phosphatase inhibitors cocktail (Abcam, Cambridge). An aliquot of the lysate was used to ascertain the protein content with a BCA protein assay kit (Pierce Biotechnology, Rockford, IL). The proteins (50 µg) were mixed with sampling buffer (no β-mercaptoethanol or heating) and separated using 10% SDS-polyacrylamide gel electrophoresis. Gel containing tyrosinase activity was successively reacted for 30 min in 100 mM sodium phosphate buffer (pH 6.8) with mild shaking. Finally, stain the gel with L-DOPA solution for 1 h.
Protease and phosphatase inhibitors cocktail
Manufactured by Abcam
Sourced in United Kingdom
Protease and phosphatase inhibitors cocktail is a mixture of small molecule inhibitors designed to prevent the degradation of proteins and phosphorylated species during sample preparation and analysis. The cocktail contains a combination of protease and phosphatase inhibitors that target a broad spectrum of enzymes to preserve the native state of the sample.
Automatically generated - may contain errors
2 protocols using protease and phosphatase inhibitors cocktail
1
Tyrosinase Zymography Assay Protocol
2
Synthesis and Characterization of MDPV
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Pure racemic MDPV • HCl was synthesized and characterized in our laboratory as previously described (Novellas et al., 2005) . Cocaine was provided by the Spanish National Institute of Toxicology. Both MDPV and cocaine solutions were prepared in 0.9% NaCl (saline, pH=7.4) immediately before administration.
The protease and phosphatase inhibitors cocktail was purchased from Abcam (Cambridge, UK). BS 3 [bis(sulfosuccinimidyl)suberate] was from ThermoFisher Scientific (Rockford, USA) and [ 3 H]WIN 35428 was from Perkin Elmer (Boston, USA). All the other reagents were of analytical grade and purchased from several commercial sources.
The protease and phosphatase inhibitors cocktail was purchased from Abcam (Cambridge, UK). BS 3 [bis(sulfosuccinimidyl)suberate] was from ThermoFisher Scientific (Rockford, USA) and [ 3 H]WIN 35428 was from Perkin Elmer (Boston, USA). All the other reagents were of analytical grade and purchased from several commercial sources.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!