RADIOIMMUNOASSAY
Introduction: –
Used to determine the concentrations of antigen present in the blood serum of the patient.
Principle:-
It is best on competitive binding between the Rio levels antigen (hot antigen) and unlebeled antigen (cold Antigen) with selected antibody and ultimately radio activities .
Radioactive material:-I125, C14 ,H3
Two method:-
Qualitative method
Quantitative Methods
1.Qualitative method:-
1.Radiolabeled antigen attached with antibody and extra antigen present in free form.
2.Wash the Micro Titre plate with a buffer Then extra antigens are removed.
Buffer:-
1%trifluoroacitic acid
60% acetonitrile + 1% TFA +39% Distilled water
3. Hot antigens are replaced by cold antigens represented by red color dots.
4.Hot antigens are washed with buffer solutions.
5.Then solution will be collected in the test and centrifuge then the supernatant liquid analyzed by radioactivity.
2. Quantitative method: –
Step 1 :- Radiolabeled antigen attach with antibody .
Step 2 :- no any hot antigen (radio labeled antigen) remove then 0% swing radioactivity.
Step 3:- it is swing 10% radioactivity because of 1ng/ml cold antigen(unlabeled antigen).
Step 4:-it is swing 20% radioactivity because of 2ng/ml cold antigen added.
– then increase radioactivity by the increasing of cold antigen.
-Take Micro Titre plate containing antibody.
– standard cold antigen solution.
– washing with the help of buffer solution and washed solution is collected in the test tube and Centrifuge at 1200 to 4000 RPM .
-After Centrifugation supernatant liquid is analysed in Radioactive counter.
