enzyme linked immunosorbent assay
DESCRIPTION
Enzyme Linked Immunosorbent Assay. ELISA Detection of Anti-HIV (This outline summarizes key points of ELISA protocol as outlined in the handout.). ANTI-HIV Simulate LABORATORY. Introduction to the protocol Laboratory Steps. Enzyme Linked Immunosorbent Assay (ELISA). - PowerPoint PPT PresentationTRANSCRIPT
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Enzyme Linked Immunosorbent Assay
ELISADetection of Anti-HIV
(This outline summarizes key points of ELISA protocol as outlined in the handout.)
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ANTI-HIV Simulate LABORATORY
Introduction to the protocol
Laboratory Steps
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• A diagnostic test for Antibodies to HIV
Enzyme Linked Immunosorbent Assay
(ELISA)
Antibodies
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Antibodies Antigens
Immune system Non-self
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Objective: to use the ELISA to detect anti-HIV from 2 donors.
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Supplies needed /group 4 transfer pipets
1 transfer
1 set of samples
1 microtiter plate (12 wells)
1 beaker of Phospate buffered saline (PBS)
1 empty beaker for “waste”
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Transfer pipets Try to approximately add the Same volume to each well.
Approximate volume 0.l ml
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Transfer Pipets for rinsing steps
Label transfer pipets
PBS 1 PBS 2 PBS 3 PBS 4
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Let’s
ELIS
A
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A. Add .1 ml of antigen to each well
Remove antigen solution
Rinse well one time with PBS
Incubate 5-10 minutes at room temperature
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B. Add 0.1 ml of PBS + control Donor 1 Donor 2
Row 1 Row 2 Row 3 Row 4
Row 1
Row 2
Row 3
Row 4
Incubate 10-15 min. at 37C
NOT IN A WATER BATH!!
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C.
Remove solutions from each well
Rinse with PBS
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D.
Add .1 ml of labeled 2nd Ab to each well
Remove 2nd antibody solution
Rinse wells with one time PBS
Incubate5-10 minutes at 37C
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E.
Add .1 ml of substrate to each well
Identify wells with anti-HIV
5-10 minutes at 37C
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OVERVIEW
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Microplate Readers
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Examples of Antibody dilutions
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Hand-In: A drawing of the steps involved in the ELISA
The results of your laboratory results
The interpretation of your results
Provide an example of how you could obtain a false positive and a false negative result.
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