WesternBlots - ImmunoBlots                             

 

What is WesternBlot?

 

Whereas ELISA measures antibody to whole virus and gives a "positive," "negative" or indeterminate test result, western blotting is a more specific test. It allows one to visualize antibodies directed against each viral or bacterial protein. For this reason, WesternBlot is a confirmatory test for a positive ELISA tests.

For example, for specific HIV diagnostic test, HIV infected cells are lysed, subjected to SDS-PAGE (sodium dodecyl sulphate polyacrilamid gel electrophoresis) and blotted onto a membrane as described above. The membrane was blocked, cut into strips and incubated with the serum samples from each patient as indicated.

Below you can see an example of a Western Blot and how to interpret it.

Viral Proteins (HIV) and Band Pattern Interpretation

Viral Proteins - HIV, like any other virus, is composed of a number of different proteins. The Western Blot positive control lane contains proteins from patient sera as well as HIV proteins. HIV positivity can therefore only be confirmed by the presence of the following types of proteins:

gp160

viral envelope precursor (env)

gp120

viral envelope protein (env) binds to CD4

p24

viral core protein (gag)

p31

Reverse Transcriptase (pol)

Band Pattern Interpretation

In 1987 the Centre for Disease Control along with several other organizations established criteria for serologic interpretation of HIV WesternBlot tests. The criteria are listed below:

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No bands present     -     Negative

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Bands at either p31 or p24 AND bands present at either gp160 or gp120     -     Positive

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Bands present, but pattern does not meet criteria for positivity     -     Indeterminate

Band pattern Interpretation

  1. Lane 1, HIV+ serum (positive control)
  2. Lane 2, HIV- serum (negative control)
  3. Lane A, Patient A negative result
  4. Lane B, Patient B indeterminate result
  5. Lane C, Patient C positive results

You can see also Helicobacter Pylori Westernblot Interpretation 

 

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