ELISA

Question 1

What does the ELISA relay on?

Answer 1

Relies on the specific interaction between the antibodies and the antigen

Question 2

What is ELISA used for?

Answer 2

Detect and is used as a diagnostic tool in medicine

Question 3

What binds to what?

Answer 3

Antigen binds to antibody

Question 4

What are the steps of the sandwich ELISA step 1-5

Answer 4

1. Plate coated with capture antibody( Specific to protein to detect)

2.Plate is incubated with capture antibody that binds to bottom of the well

3.After incubation the liquid is removed into waste beaker and pat dry with tissue

4.Use PBS-T or TBS-T to wash/remove the unbound material in the wells x3-5 times - pour into waste beaker

5.Add blocking buffer to the wells (Mixture of proteins) to prevent the binding of proteins to areas of the wall not occupied by capture antibody

Question 5

What are the steps of the sandwich ELISA step 6-9

Answer 5

6. After block buffer it is washed oft remove unbounded proteins x3-5 times wash

7.Add sample (Crude mixture of proteins

8.After incubation the capture antibody binds to protein of interest then use was buffer to remove unbounded

9.Then probe the presence of the antigen using the antibodies

Question 6

What are the steps of the sandwich ELISA step 10-14

Answer 6

10. Primary antibody is added to detected antigen
11. then washout with each buffer

12.Add secondary antibody, bound to an enzyme to recognise the primary antibody.

13.Sa brings substrate, then sa is washed off

14. then incubate and add the enzyme substrate

Question 7

What is the order of the sandwich ELISA

Answer 7

1. Capture antibody
   2.Blocking Buffer
   3.Sample
   4.Primary Antibody
   5.Secondary Antibody
2. Enzyme substrate

Question 8

How does direct ELISA work?

Answer 8

1. Antigen coating
   2.wash
   3.blocking buffer
   4.wash
2. Primary antibody
3. wash
4. enzyme causing substrate to change colour and it will break detected.

Question 9

How does competitive ELISA work?

Answer 9

1. Coating with antigen
2. Blocka buffer
   3.remove and wash add sample
3. wash then Add primary and wash
   5.Add inhibitor antibody with enzyme
4. Wash and let the enzyme oxidase the substrate and generate a colour

Question 10

What are examples of Antigens?

Answer 10

Virus
Bacteria
Proteins

Question 11

What can ELISA be used for?

Answer 11

Bacterial infection such as Lyme disease
Viral infection such as HIV and Hepatitis A, B and C or flu/ covid
Fungal infection- Yeast infection