Design an ELISA experiment (STEP BY STEP) explaining how you would test the reac
ID: 3165557 • Letter: D
Question
Design an ELISA experiment (STEP BY STEP) explaining how you would test the reactivity of Ig1, Ig2, Ig3, Ig4 against antigen AIP-2 (anti-inflammatory 2) of hookworm. Then design a mouse model experiment to test the same reactivity of the named IgGs against AIP-2.
Background: Hookworm is known to secrete the anti-inflammatory protein 2. IgG1 can be used to test the presence of antigen in blood. Ig2 and Ig3 can be used as control while IgG4 is known to be produced in the body by the hookworm antigen
Explanation / Answer
ELISA Experiment STEP by STEP Process to test reactivity of Ig1, Ig2, Ig3, Ig4 with antigen AIP-2 in hookworm
1. Antibody Coating
Specific capture antibody of Ig1, Ig2, Ig3, Ig4 is immobilized on high protein-binding plates by overnight incubation. Plates are blocked with irrelevant protein e.g. albumin.
2. Protein Capture
Samples of AIP-2 (Auto Immune Pancreatitis) and standard dilutions are added to the wells and will be captured by the bound antibodies (Ig1)
3. Detection of Antibody
Specific Ig1, 2, 3, 4 detection antibody is added to the wells to enable detection of the captured protein.
4.Enzyme Conjugate
Enzyme conjugated with substrates is added to the wells and will bind to the Ig1,2 3, 4 antibody.
5. Addition of Substrate
Colorimetric substrate is added to the wells and will form a colored solution when catalyzed by the enzyme.
Design a mouse model experiment to test the same reactivity of the named IgGs against AIP-2
1. Antibody Coating
Specific capture antibody of IgGs is immobilized on high protein-binding plates by overnight incubation. Plates are blocked with irrelevant protein.
2. Protein Capture
Samples of AIP-2 (Auto Immune Pancreatitis) and standard dilutions are added to the wells and will be captured by the bound antibodies (IgGs)
3. Detection of Antibody
Specific IgGs detection antibody is added to the wells to enable detection of the captured protein.
4.Enzyme Conjugate
Enzyme conjugated with substrates is added to the wells and will bind to the IgGs antibody.
5. Addition of Substrate
Colorimetric substrate is added to the wells and will form a colored solution when catalyzed by the enzyme.
6. Finally, In hookworm, high affinity will be showing for mouse model when you analyse with IgGs against with AIP-2.
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