Enzyme II: medical uses of enzymes

Question 1

ELISA: enzyme-linked immunosorbent assay is used too is used to detect what diseases?

Answer 1

Detection of Mycobacterium antibodies in tuberculosis
Detection of hepatitis B markers in serum
Detection of enterotoxin of E. coli in faeces
Detection of HIV antibodies in blood samples
Hormone levels

Question 2

1st step of ELISA : 96-well plate

Answer 2

Virus proteins (antigens) are added
to wells of a 96-well plate.

Question 3

2nd step of ELISA: antigen binding

Answer 3

The antigens bind to the plastic,

coating the bottom of the wells

Question 4

3rd step of ELISA: primary antibody

Answer 4

The primary antibody is added to the well.
In the case of the dengue ELISA,
the primary antibodies (IgM) are from
the patient’s serum sample

Question 5

4th step of ELISA : washing

Answer 5

Excess antibody is washed away, leaving
only antibodies bound to the antigens
behind. This wash removes excess
antibodies that are unbound and prevents
non-specific binding.

Question 6

5th step of ELISA: secondary

Answer 6

A (HRP conjugated antibody) secondary antibody is added to the wells.
This antibody recognises the patient
IgM antibodies, bound to the antigens.
The secondary antibody also has a
colorimetric tag attached.

Question 7

6th step of ELISA: washing

Answer 7

Excess secondary antibody is washed away,
leaving only secondary antibodies,
bound to the patient IgM antibodies.
This wash removes excess antibodies
that are unbound.

Question 8

7th step of ELISA: substrate addition

Answer 8

A HRP substrate is added to the wells.

Question 9

8th step of ELISA(last step ) : colour arise

Answer 9

Bound secondary antibody containing a
colorimetric tag will cause a colour change
when exposed to the HRP substrate. A colour
change indicates a positive reaction.