Topic 3.2: Protein Detection & Analysis
You are about to begin Topic 2, Protein Detection & Analysis.
In this topic you’ll learn about the role of Protein Detection & Analysis. You will start by clicking on the next button at the bottom of the page to being reviewing information about Protein Detection & Analysis.
Upon completion of this module topic, you should:
- Be able to use electrophoresis techniques to analyze, purify and identify proteins by using the technique to separate proteins according to their electrophoretic mobility to estimate their molecular weights.
- Be able to prepare a sodium dodecyl sulfate (SDS) polyacrylamide gel (PAG) for protein electrophoresis (E). (SDS-PAGE)
- Be able to prepare protein samples from tissue and cell samples for electrophoresis (SDS-PAGE) using protein extraction, homogenization, denaturation and preparation techniques.
- Be able to assemble to SDS-PAGE apparatus for running a protein gel.
- Be able to load prepared protein samples on to a polyacrylamide gel for electrophoresis.
- Be able to run SDS-PAGE using electrical current generated through a power supply.
- Be able to conduct the Western Blot technique following SDS-PAGE to detect specific proteins in samples of tissue homogenates or extracts.
- After conducting SDS-PAGE, be able to transfer separated proteins to a nitrocellulose or PVDF membrane by appropriately assembling the Western Blot transfer apparatus and preparing (equilibrating and charging) the transfer membrane.
- Be able to probe the proteins transferred to the nitrocellulose or PVDF membrane using antibodies specific to the target protein (primary antibody specific to the target protein and a secondary antibody containing a detection signal such as horseradish peroxidase specific to the primary antibody bound to the target protein).
- Be able to analyze the probed transfer membrane using Ultraviolet fluorescence or gel staining techniques to display your probed protein target(s).
- Be able to interpret your Western Blot results by estimating the size(s) of your detected protein(s) from your samples.
- Be able to explain and perform an Enzyme-Linked Immunosorbant Assay (ELISA) to detect the presence of a particular antibody or an antigen in a sample.
- Understand the mechanisms of the ELISA technique and its important applications to medicine and disease diagnosis.
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