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Topic 3.2: Protein Detection & Analysis

Screenshot of videoYou 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.

OBJECTIVES:

Upon completion of this module topic, you should:

  1. 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.
  2. Be able to prepare a sodium dodecyl sulfate (SDS) polyacrylamide gel (PAG) for protein electrophoresis (E). (SDS-PAGE)
  3. Be able to prepare protein samples from tissue and cell samples for electrophoresis (SDS-PAGE) using protein extraction, homogenization, denaturation and preparation techniques.
  4. Be able to assemble to SDS-PAGE apparatus for running a protein gel.
  5. Be able to load prepared protein samples on to a polyacrylamide gel for electrophoresis.
  6. Be able to run SDS-PAGE using electrical current generated through a power supply.
  7. Be able to conduct the Western Blot technique following SDS-PAGE to detect specific proteins in samples of tissue homogenates or extracts.
  8. 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.
  9. 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).
  10. Be able to analyze the probed transfer membrane using Ultraviolet fluorescence or gel staining techniques to display your probed protein target(s).
  11. Be able to interpret your Western Blot results by estimating the size(s) of your detected protein(s) from your samples.
  12. 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.
  13. Understand the mechanisms of the ELISA technique and its important applications to medicine and disease diagnosis.

 

 
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