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SKU AR1182
Pack size 1 mL
Form Solution (liquid)
Applications Western blotting, protein purification, Co-IP; assays for protein expression, activity, modification, profiling and characterization, quantitative measurement; epitope tagging; reporter gene analysis
Price: $50.00
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Overview

Product Name Broad Spectrum Protease Inhibitor Cocktail
Synonyms Broad Spectrum Protease Inhibitors
Description Broad Spectrum Protease Inhibitor Cocktail (100X) is a Western blot related ready-to-use concentrated stock solution reagent containing a blend of seven protease inhibitors that is to be added to cell lysis buffer to protect the integrity and functionality of native cellular proteins against degradation by multiple classes of endogenous proteases during protein extraction and sample preparation procedures.
Application Western blotting, protein purification, Co-IP; assays for protein expression, activity, modification, profiling and characterization, quantitative measurement; epitope tagging; reporter gene analysis
Pack Size 1 mL
Reagent Type Western Blotting related reagent; Inhibitors
Usage Protect native cellular proteins from destructive degradation by endogenous proteases following cell lysis
Preserve native cellular proteins intact and functional
Screen extracts for proteolytic activity
Study proteolysis in the regulation of cellular processes
Content E-64, AEBSF-HCL, bestatin, leupeptin, aprotinin, Pepstatin, and EDTA stabilized in dimethylsulfoxide (DMSO)
Target Specificity Serine proteases, cysteine proteases, aspartic acid proteases, metalloproteases, aminopeptidases
Target Sample Cell lysis extracts

BioChemical Properties

Protease Inhibitor Component MW Protease Family Targeted Inhibition Type Typical Working (1X) Conc. Concentration in 100X cocktail
E-64 357.4 Cysteine proteases (papain, calpain, lysosomal cathepsins) Irreversible 1 to 20µM 1.5mM
AEBSF-HCI 239.5 Serine proteases (trypsin, chymotrypsin, plasmin, trypsinogen, urokinase, kallikrein) Irreversible 0.2 to 1.0mM 100mM
Bestatin 308.4 Amino-peptidases Reversible 1 to 10µM 5mM
Leupeptin 475.6 Serine and cysteine proteases Reversible 10 to 100µM 2mM
Aprotinin 6511.5 Serine proteases (trypsin, chymotrypsin, plasmin, trypsinogen, urokinase, kallikrein) Reversible 100 to 200nM 80µM
Pepstatin A 685.9 Aspartic acid proteases (pepsin and rennin) Reversible 1 to 20µM 1mM
EDTA 372.2 Metalloproteases (thermolysin and carboxypeptidase A) Reversible 2 to 10mM 2mM

Properties

Form Supplied 100X concentrated stock solution
Physical State Liquid, colorless, transparent
Pack Size 1 ml
Recommended working concentration 100-fold dilution in tissue lysis buffer
10 µl of the Protease Inhibitor Cocktail solution is enough to inhibit degradation of proteins in 1 ml lysate
Compatibility with reagents Fully compatible with cell lysis buffers and Broad Spectrum Phosphatase Inhibitor Cocktail
Compatibility with assays Not MS-compatible: contain AEBSF;
Not compatible with immobilized metal chelate affinity chromatography and 2D gel electrophoresis: contain EDTA
Storage Store at -20˚C for one year
Safety Precautions Harmful. PMSF is toxic and causes irritation to the eyes and skin. Please operate with caution and wear eye and hand protection and proper lab garments. FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC AND CLINICAL USE

Usage and Handling

The product is supplied as a 100X concentrated stock solution in a liquid format for improved accuracy, solubility, and ease of use in comparison to traditional tablets.
1. Add broad spectrum protease inhibitor cocktail in lysis buffer in the ratio of 1:100 and mix well.
2. Add the solution in to samples for protein extraction.
Since endogenous protease levels may vary among cell and tissue types, it is up to the researcher to optimize the concentration for their experiments.

Background

Crude cell extracts contain a number of endogenous enzymes, such as proteases and phosphatases, which are capable of digesting the proteins present in the extract. An optimized method to improve the yield of intact and functional native proteins is to add inhibitors of these enzymes known to be present in the source material. This broad spectrum protease inhibitor cocktail is a complex of various protease inhibitors, which has been tested for inhibiting proteases and esterase broadly.
Protease inhibitors are critical reagents for the preservation of protein integrity during purification and analysis procedures by knocking out specific proteases to avoid peptide bond hydrolysis and subsequent protein destruction. They are biological or chemical compounds that function by reversibly or irreversibly binding to the protease. Most known proteases belong to one of four evolutionarily distinct enzyme families based on the functional groups involved in the peptide bond cleavage. No single chemical is effective for all types of proteases. A mixture or "cocktail" of several different inhibitor compounds is commonly used that consistently inhibit a multitude of protease classes to ensure that protein extracts do not degrade before analysis for target proteins of interest. Not using protease inhibitors leads to the loss of a large number of valuable proteins in a lysate sample, adversely affecting downstream applications by biologically meaningless representation of protein activities and gaining false negative immunostaining results for targets of interest.

Broad Spectrum Protease Inhibitor Cocktail (100x) Images

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Broad Spectrum Protease Inhibitor Cocktail (100x)
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Publications

Liang, Y., Song, D.Z., Liang, S., Zhang, Z.F., Gao, L.X., & Fan, X.H. (2017). The hemagglutinin-neuramidinase protein of Newcastle disease virus upregulates expression of the TRAIL gene in murine natural killer cells through the activation of Syk and NF-κB. PLoS One, 12(6), e0178746. doi: 10.1371/journal.pone.0178746