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Alzheimer's Disease | AssayBiotechnology
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Alzheimer's Disease

This How To Guide Will Cover

  • Western Blot Theory Overview
  • Sample Preparation
  • SDS-PAGE Method
  • Transferring Method
  • Immunoblotting Method
  • Detection Method

Western Blot Theory Overview

Antibodies bind to specific sequences of amino acids, known as the epitope. Because amino acid sequences are different from protein to protein, antibodies can recognize specific proteins among a group of many. Therefore, a single protein can be identified in a cell lysate that contains thousands of different proteins and its abundance quantified through western blot analysis. First, proteins are separated from each other based on their size. Second, antibodies are used to detect the protein of interest. Finally, a substrate that reacts with an enzyme is used to view the antibody/protein complex.

Sample Preparation

The first step in sample preparation, is isolating the protein from a source. Usually, protein is purified from cells. However, other sources of protein, such as synthetically derived protein, can also be run through a Western blot.

     

    Next, the protein concentration is determined. Loading buffer, which contains SDS (sodium dodecyl sulfate), and BME (beta-mercaptoethanol) is added to the protein suspension.

     

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