HIV Research / Invitro Studies

In invitro lab studies, this drug was put for Drug Sensitivity Test and Anti-retroviral Activity Test in the laboratory adhering to the standard protocol and the highest inhibition rate of 98% was recorded against the causative agent, HIV virus. The detail procedures undertaken in an invitro lab study are mentioned as under :-

Procedure of the invitro lab study

1. Collect 20 ml Blood in an EDTA containing vial
2. Mix it properly and keep at room temperature
3. Overlay this blood sample on 2 ml Histopaque solution in 25-ml sterile centrifuge tube. Centrifuge the tube at 1500 rpm for 20 min
4. Two layers will be formed after centrifugation
5. Separate out the upper layer of Histopaque with the help of sterile pipette and collect the lymphocyte ring in 2 ml sterile eppendorf tube
6. Microfuge it for 1 min at the bottom of the eppendorf tube lymphocytes sediment will be observed.
7. Re-suspend the sediment in 2-ml fresh RPMI-C Media.
8. Prepare the serial dilutions of the drug of our interest or to be tested in the range of 1ug/ml - 500 ug/ml
9. Add 500 ul of the lymphocyte culture with the help of sterile pipette / tips and add the appropriate dilution of the drug to the respective culture well along with control without addition of any drug.
10. Incubate it in CO2 incubator at 37 degree Celsius (5% CO2)
11. After incubation do the P-24 ELISA test for Control and for Test
12. Interpret the results on the basis of difference in between the Control O.D and Test O.D and calculate the % Inhibition of the viral growth in Test in comparison with Control.

Example: - O.D of the Control always ranges from 0.2-0.4 while the O.D of Test ranges in between 0.06-0.1, which indicates the inhibition of the virus multiplication.