Biology Forums - Study Force

What is the difference between the Western blot and  immunocytochemistry. What are the benefits and drawbacks of each?

In a Western Blot, a protein sample is run on an acrylamide gel, and then transferred to a fixed membrane. The membrane can then be washed and stained with a antibody to a specific protein of interest. This will reveal whether or not the protein of interest is present in the sample or not and it's approximate size and quantity.

Immunocytochemistry is another laboratory technique that uses antibodies that target specific peptides or protein antigens in the cell via specific epitopes (a site on an antigen at which an antibody can bind, the molecular arrangement of the site determining the specific combining antibody). These bound antibodies can then be detected using several different methods, such as ELISA (see the animation link below). Immunocytochemistry allows researchers to evaluate whether or not cells in a particular sample express the antigen in question. In cases where an immunopositive signal is found. Immunocytochemistry also allows researchers to determine which sub-cellular compartments are expressing the antigen.

In immunohistochemistry, tissues are sliced and prepared on microscope slides, fixed and treated with an antibody of interest that can reveal the presence of a protein of interest and it's location in or on a cell for example.

Check out this tutorial: http://www.ppdictionary.com/tutorials/elisa.htm (http://www.ppdictionary.com/tutorials/elisa.htm) (Everything will make sense).

For Immunocytochemistry, antibodies generated to a specific protein can be used to find that unique protein in a cellular extract. Traditionally scientists used chromatography media to secure the antibodies to a solid support. However, a chromatography column prevented researchers from handling many samples. Low-pressure columns could only be run so fast. So the research community sought a technique that could process more samples. The ELISA technique takes smaller volumes and hence lesser amounts of reagents.

To detect or visualize the antigen-antibody complex, the antibody must be labeled. In the past, researchers had to conjugate their own antibodies and labels. Now, though, they can routinely buy antibodies conjugated to many different labels. The labels include radioisotopes; horseradish peroxidase, alkaline phosphatase, and other enzymes; fluorescent tags such as fluorescein; and biotin.

Rapid tests are flexible, time to get  results is less than 30 minutes. Skilled staff is not required. Results are very easy to interpret and permit on-site testing. ELISAs are good for high volumes of specimen. Not that expensive to perform tests.

The Western Blot is the most common test use for confirmation of HIV infection.  However western blot tests have several Disadvantages such as lack of standardization in performance and interpretation. They give a high range of indeterminate results, they are complex, in addition they are very expensive.

Reply back if you need further assistance.

Thank you for your response.. It was very helpful! :)