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W19- Microbiology & Virology > ELISA > Flashcards

Flashcards in ELISA Deck (12)
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1
Q

What does ELISA stand for and what is it

A
  • Enzyme-linked immunosorbent assay

- A technique used to detect (assay) specific molecules in samples

2
Q

What are the 3 ways in which ELISA is used in veterinary medicine

A
  • In disease detection
  • In detection of illegal drugs
  • Detection of hormones
3
Q

What is indirect ELISA used for

A
  • used to detect/measure a specific antibody in a sample
4
Q

How does an indirect ELISA work

A
  1. Antigen bound to plastic well
  2. Well is coated with blocking reagent to prevent non-specific binding and washed to remove excess reagent
  3. Test sample is added and any antiodies in that sample that react against antigen will bind. Wash to remove excess
  4. Secondary antibody with attached enzyme added. Wash to remove excess
  5. Substrate is added
  6. Substrate converted by enzyme if antibody is present
5
Q

What is direct ELISA used for

A

used to detect / measure the concentration of specific antigens in a sample

6
Q

What are the 2 types of direct ELISA

A
  • Sandwich ELISA

- Competitive ELISA

7
Q

How does a sandwich ELISA work

A
  1. Wells coated with capture antibody
  2. Well is coated with blocking reagent to prevent non-specific binding and washed to remove excess reagent
  3. Test sample is added and any antigen can bind to capture antibody. Wash to remove excess
  4. Addition of primary antibody results in sandwich formation. Wash to remove excess
  5. Add secondary antibody with attached enzyme. Wash to remove excess
  6. Substrate is added and converted by enzyme if antigen present
8
Q

When is competitive ELISA used instead of sandwich ELISA

A

If the antigen is very small and has only one epitope (binding site for antibody)

9
Q

What does a positive control in ELISA allow for

A

Confidence in the diagnostic test results, confirms that the negative results are accurate and assists in any protocol adjustments or optimizations that may be necessary

10
Q

What does a negative control in ELISA show

A

If any solutions of the assay are contaminated adding validity to any positive results

11
Q

In a sandwich ELISA what would happen if the blocking step was ommitted

A

All wells including the negative control wells, would show uniform over development

12
Q

In a sandwich ELISA what would happen if there was no washing steps after the detection antibody was added to the wells

A

All wells, including the negative control wells would show uniform over development