The microtitration plate wells are coated with HIV-1 and HIV-2
recombinant antigens. The sample is incubated in the wells.
If sample contains antibodies against HIV-1 or HIV-2 they will
bind to the antigens present in the wells. The unbound fraction is eliminated by washing. In the next step, is added the
conjugate which contains the same antigens than the plate
conjugated to peroxidase. They will bind to the antibodies if
they were present in the sample. The unbound conjugate is
removed by washing.
Then, a solution containing tetramethylbenzidine (TMB) and
hydrogen peroxide is added. The reactive samples develop
a light blue color that changes to yellow when the reaction
is stopped with sulfuric acid.
Coated microtitration plate: microtitration plate with removable strips and 96 wells, coated with HIV-1 and HIV-2
Conjugate: recombinant antigens bound to peroxidase.
Substrate: tetramethylbenzidine (TMB) and hydrogen peroxide solution.
Stopper: 2 N sulfuric acid.
Concentrated Wash Buffer: saline buffer with surfactant
agent (25x). Green color.
Positive Control: inactivated human serum containing antiHIV-1 and anti-HIV-2 antibodies. Orange color.
Negative Control: inactivated human serum, non-reactive.
Distilled or deionized water.
NON-PROVIDED REQUIRED MATERIAL
- Micropipettes for measuring stated volumes
- Disposable tips
- Volumetric material for stated dilutions
- Incubator at 37ºC
- Disposable gloves
- Absorbent paper
- Alarm clock or stopwatch
- Sodium hypochlorite
- (Automatic or manual) microtitration plate wash system
- Spectrophotometer for microtitration plate reading