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Analysis of carbonic anhydrase by Differential Scanning Fluorimetry(DSF) (CAT#: STEM-MB-0758-WXH)

Introduction

Carbonic anhydrase is an enzyme that assists rapid inter-conversion of carbon dioxide and water into carbonic acid, protons and bicarbonate ions. This enzyme was first identified in 1933, in red blood cells of cows. Since then, it has been found to be abundant in all mammalian tissues, plants, algae and bacteria.




Principle

Differential Scanning Fluorimetry measures protein thermal unfolding by monitoring changes in fluorescence emission of a sample upon heating. This allows the determination of protein thermostability and complex formation even with weakly binding ligands by thermal shift assay. Differential Scanning Fluorimetry is therefore ideally suited for screening of optimum buffer conditions like pH, buffer composition and ionic strength. The technique is applicable to any biological sample, from soluble proteins to integral membrane proteins.

Applications

To identify low-molecular-weight ligands that bind and stabilize purified proteins.
To measure the denaturation and unfolding of proteins.

Procedure

1. Preparation of compound solutions
2. Preparation of buffer/additive screen plates
3. Preparation of compound storage plates
4. Equipment preparation
5. Sample preparation
12. Performing the scan

Materials

Real-time PCR instrument