(Download) "Highly Sensitive Immunoprecipitation Method for Extracting and Concentrating Low-Abundance Proteins from Human Serum (Technical Briefs)" by Clinical Chemistry ~ eBook PDF Kindle ePub Free
eBook details
- Title: Highly Sensitive Immunoprecipitation Method for Extracting and Concentrating Low-Abundance Proteins from Human Serum (Technical Briefs)
- Author : Clinical Chemistry
- Release Date : January 01, 2005
- Genre: Chemistry,Books,Science & Nature,
- Pages : * pages
- Size : 233 KB
Description
Purification and identification of small, low-abundance proteins from complex samples such as serum for use in Western blotting experiments has long been a challenge. Unfortunately, commercially available methods are not always suitable for small sample sizes and low-abundance proteins such as cardiac troponin T (cTnT). In this report we present a highly sensitive immunoprecipitation method we have developed for extracting and concentrating low-abundance proteins from serum. We compared the performance of our method with that of the commercially available ImmunoPure[R] Protein-A IgG orientation Kit (Pierce), using antibodies directed against cTnT and serum of a patient with acute myocardial infarction (AMI). Both methods are based on the binding of antibodies to Sepharose beads and subsequent cross-linking with a diimido ester. The formation of amidines between imido esters and amines from protein side chains was first described in 1962 by Hunter et al. (1), who suggested this for cross-linking purposes. Levy et al. (2) described the use of diimido esters for covalent coupling of antibodies to immobilized antigens. Later studies reported similar methods for antibody immobilization with cross-linkers such as dimethyl pimelimidate and dimethyl suberimidate (3, 4).