MICRO Module 7

Question 1

What does the host immune system do upon exposure to microbial antigens?

Answer 1

The host immune system makes specific antibodies to that antigen to combat the infection.

Question 2

What occurs when an antibody reacts with a soluble antigen?

Answer 2

A precipitation reaction will occur.

Question 3

Why are precipitation reactions important?

Answer 3

They are important indicators of disease in patients.

Question 4

What happens to antibody concentration during the course of a disease?

Answer 4

The antibody concentration in the patient’s blood will increase.

Question 5

What can the presence of antibodies to specific pathogens indicate?

Answer 5

It can serve as an indicator of the course of an infection.

Question 6

What is the serum titer?

Answer 6

The highest dilution of the serum that can still precipitate the antigen.

Question 7

What does an increasing titer over time indicate?

Answer 7

It is indicative of an active infection.

Question 8

What is the overall learning objective of the Gel Immunodiffusion Protocol?

Answer 8

Describe and analyze the specific interactions between antigen and antibody using gel immunodiffusion assay.

Question 9

What are the detailed learning objectives of the Gel Immunodiffusion Protocol?

Answer 9

1. Describe the principle of immunodiffusion assay.
2. Analyze the results of immunodiffusion assay and the applications of this assay.
3. Predict expected results of an experiment.
4. Use controls for an immunodiffusion assay.
5. Analyze the results of an experiment, including how they relate to the applications of this assay.

Question 10

What is the purpose of the Gel Immunodiffusion assay?

Answer 10

To study the specificity of the antigen and antibody reaction and understand its diagnostic applications.

Question 11

What are antigens?

Answer 11

Foreign substances that elicit a reaction from the immune system of the host, such as chemicals, pollen, or proteins on microbes.

Question 12

What are antibodies?

Answer 12

Proteins produced by the host’s immune system in response to exposure to antigens.

Question 13

How do antibodies interact with antigens?

Answer 13

Antibodies recognize and bind to antigens with high specificity, helping to eliminate the antigen from the body.

Question 14

How can specific antigen-antibody interactions be detected?

Answer 14

By various methods, including the gel immunodiffusion assay, which forms insoluble precipitates detectable on a gel-like matrix.

Question 15

What is the typical setup for a gel immunodiffusion assay?

Answer 15

A series of wells are cut in an agarose gel, with antigen in the center and antibodies in peripheral wells.

Question 16

What indicates a positive reaction in a gel immunodiffusion assay?

Answer 16

The formation of a white precipitate at the zone where optimal concentrations of antibody and antigen meet.

Question 17

What is a positive control in the gel immunodiffusion assay?

Answer 17

Serum from a sample known to contain antibodies specific to the antigen, resulting in a white precipitate.

Question 18

What is a negative control in the gel immunodiffusion assay?

Answer 18

Serum that does not contain antibodies or just the buffer, which should not yield a positive reaction.

Question 19

How is the titer of antibodies estimated in the assay?

Answer 19

By making serial dilutions of the patient sample and determining the highest dilution at which antibodies can be detected.

Question 20

What is the diagnostic application of the gel immunodiffusion assay?

Answer 20

It can be used as a preliminary tool for diagnosing infections, particularly in veterinary sciences.

Question 21

What is the first step in the Gel Immunodiffusion Protocol?

Answer 21

Obtain an agarose gel plate with premade wells and label the bottom of the plate.

Question 22

What dilutions should be made of the patient’s sample?

Answer 22

Undiluted, 1:2, 1:4, 1:8, 1:16, 1:32.

Question 23

How much antigen is placed in the central well?

Answer 23

0.010 mL of antigen.

Question 24

What should be done after adding the samples to the wells?

Answer 24

Wrap the plate in plastic wrap and place it at room temperature.

Question 25

How long should the gel immunodiffusion assay be incubated?

Answer 25

For 6-8 hours.

Question 26

What should be recorded after incubation?

Answer 26

The highest dilution at which a precipitate appears.

Question 27

What does ELISA stand for?

Answer 27

Enzyme-linked Immunosorbent Assay

Question 28

What is the primary use of ELISA?

Answer 28

To detect unknown antibodies or antigens in a patient sample.

Question 29

Which viral infections can be detected using ELISA?

Answer 29

Influenza, HIV, and Hepatitis B or C.

Question 30

Can ELISA be used to detect bacterial infections?

Answer 30

Yes, it can detect bacterial infections like Salmonellosis and Strep-throat.

Question 31

What is the overall learning objective of the ELISA protocol?

Answer 31

Describe and analyze the specific interactions between antigen and antibody using ELISA.

Question 32

What is the first step in a typical ELISA experiment?

Answer 32

A known, pure antigen is linked to the wells of a plastic plate.

Question 33

What happens after the known antigen is linked to the plate?

Answer 33

Excess unbound antigen is washed off.

Question 34

What is added to the plate after the unknown sample is incubated?

Answer 34

A secondary antibody that is linked to an enzyme.

Question 35

What indicates the presence of antibodies in the unknown sample?

Answer 35

The development of a colored product, typically blue.

Question 36

How is the quantity of antibodies estimated in an unknown sample?

Answer 36

By comparing the intensity of the color to known standards.

Question 37

What is the purpose of including controls in an ELISA experiment?

Answer 37

To validate the results of the experiment.

Question 38

What is the expected result for a positive control in ELISA?

Answer 38

Development of blue color.

Question 39

What is the expected result for a negative control in ELISA?

Answer 39

Absence of blue color.

Question 40

What was detected in patients X and Y using ELISA?

Answer 40

The presence of antibodies to influenza antigen HA.

Question 41

What distinguishes patient X from patient Y in the context of the ELISA results?

Answer 41

Patient X had influenza virus a couple of months ago, while patient Y has never had influenza.

Question 42

Where can videos on Quantitative and Qualitative ELISA be found?

Answer 42

On YouTube under Module 4 learning materials.

Question 43

What is the principle behind ELISA?

Answer 43

ELISA (Enzyme-Linked Immunosorbent Assay) is a test that detects the presence of antibodies in a sample by using an antigen immobilized on a plate and a substrate that produces a measurable signal.

Question 44

Interpret the results for the 5 patients based on the ELISA data. Which patients are positive/negative for antibodies to Borrelia burgdorferi?

Answer 44

Patients A, B, and C are positive for antibodies to Borrelia burgdorferi, while patients D and E are negative. The titer of antibodies is the highest dilution at which a positive result is observed.

Question 45

Which of these patient(s) is at a higher risk of contracting Lyme disease in the future?

Answer 45

Patients A, B, and C are at a higher risk of contracting Lyme disease in the future due to their positive antibody results.

Question 46

What do the controls in rows 6 and 7 tell you?

Answer 46

The controls in rows 6 and 7 validate the ELISA test results. If Row 7 (columns 1 & 2) showed yellow color, it would indicate a positive control, confirming that the assay is functioning correctly.

Question 47

List the steps in the ELISA setup to study the antibody response to COVID-19.

Answer 47

1. Coat the plates with purified COVID-19 S-protein antigen. 2. Incubate the plates to allow binding. 3. Add blood samples from patient volunteers who received the vaccine and placebo. 4. Incubate again. 5. Wash the plates to remove unbound antibodies. 6. Add necessary secondary antibodies conjugated to enzyme. 7. Incubate the plates. 8. Wash the plates again. 9. Add substrate for color development. 10. Use the ELISA plate reader to monitor color development.