How they work

A monoclonal antibody to HIV p24 along with HIV-1 and HIV-2 antigens are immobilised in a series of wells. The sample to be analysed is added to the well and any corresponding antibody and/or antigen is captured to form an antibody-antigen complex.

A wash step may be used at this point to remove any unbound molecules. An anti-p24 antibody and HIV-1 and HIV-2 antigens, all labelled with an enzyme or in some cases a co-enzyme, which bind to form an antibody-antigen-antibody/enzyme and/or an antigen-antibody-antigen/enzyme conjugate complex, are added and followed by a wash step. A substrate solution is added which will produce a colour change which is proportional to the amount of bound enzyme.

Thus, samples which do not contain HIV antigen and/or antibody will not form a complex and therefore no colour reaction will take place. Wells that contain samples that do contain HIV antigen and/or antibody will show a colour change corresponding to the number of individual complexes formed.

The colour produced should be measured on a spectrophotometer to give a numerical reading which will allow comparison with the assay controls.

A number of variations of the ELISA technique may be employed; the indirect ELISA is normally used to detect antibody, the antibody capture assay is used to determine the class (e.g. IgM) of a particular antibody and the competitive ELISA is usually used for antibody detection whereby an absence of colour indicates a positive result.¹