Mechanisms/Applications of Ag/Ab Assays

Question 1

Noncompetitive solid-phase EIA for Ab detection

- Reagents and equipment

Answer 1

• Ag-coated tube
• Abs in patient’s serum
• Enzyme-labeled antihuman IgG
• Substrate solution

Question 2

Noncompetitive solid-phase EIA for Ab detection

- List the steps

Answer 2

1. Abs in patient’s serum is added to an Ag-coated tube
2. Incubate and rinse
3. Add enzyme-labeled antihuman IgG and incubate and rinse
4. Add substrate solution

Question 3

Noncompetitive solid-phase EIA for Ab detection

- Interpretation and appearance of a positive and negative result

Answer 3

Pos: color change
Neg: no color change

Question 4

Competitive solid-phase EIA for Ab detection

- Reagents and equipment

Answer 4

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Question 5

Competitive solid-phase EIA for Ab detection

- List the steps

Answer 5

1. Put both Abs in patient’s serum and enzyme-labeled Abs into the Ag-coated tube
2. Incubate and rinse
3. If Ab was present → no color change
4. If Ab was absent → color change

Question 6

Competitive solid-phase EIA for Ab detection

- Interpretation and appearance of a positive and negative result

Answer 6

Pos = Ab was absent, enzymes will bind, substrate added = color change
Neg = Ab was present still bound to tube, substrate added = no color change

Question 7

Noncompetitive solid-phase EIA for Ag detection

- Reagents and equipment

Answer 7

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Question 8

Noncompetitive solid-phase EIA for Ag detection

- List the steps

Answer 8

1. Add viral Ag to an Ab-coated tube
2. Incubate and rinse
3. Add enzyme-labeled antiviral Ab and incubate and rinse
4. Add substrate solution

Question 9

Noncompetitive solid-phase EIA for Ag detection

- Interpretation and appearance of a positive and negative result

Answer 9

Pos = color change = Ag was bound

Question 10

Advantages of EIA

Answer 10

• Well standardized
• Good for large volume testing
• Can be automated or semi-automated
• Cost effective
• Requires less technical skill than many methods

Question 11

Disadvantages of EIA

Answer 11

• Unable to distinguish specific from non-specific binding

- May require repeat and/or confirmatory testing

Question 12

BioMerieux Vidas

- General format

Answer 12

Automated EIA = walk-away function

- Pipette dips into reagent wells, both Ag and Ab detection methods are available

Question 13

BioMerieux Vidas

- Solid phase

Answer 13

Pipette tip (coated on the inside)

Question 14

Microwell-based EIA instruments

- General format

Answer 14

Automated EIA = completely or largely walk away

- Used for detection of unknown Abs

Question 15

Microwell-based EIA instruments

- Solid phase

Answer 15

Wall of microwell

Question 16

Multiplex Micro-bead systems

- Clinical application

Answer 16

• Designed to detect and distinguish many different antibodies simultaneously in a single test well

Question 17

Multiplex Micro-bead systems

- Format

Answer 17

• Microbes dyed various shades of colors; each color is coated with particular antigen
• Laser interrogation; one laser identifies among of patients antibody bound and the other identifies the color