Immunological Testing

Proliferation Assays

Binding of antigen by T cells through the T-cell receptor triggers activation and proliferation. This response is antigen-specific, and requires the participation of appropriate antigen-presenting cells such as macrophages.

For example, if PBMC from an individual who has received a tetanus shot are cultured together with tetanus antigen, the small number of tetanus-specific cells will begin dividing and secreting cytokines. Since the number of cells that proliferate is too small to detect by counting the cells, proliferation is usually detected by measuring the incorporation of 3H-thymidine (thymidine is incorporated into the DNA of dividing cells).

The amount of 3H-thymidine incorporated in response to antigen stimulation is compared to incorporation in control wells to yield a stimulation index (SI). In general, SI's greater than 3 are considered evidence of an antigen-specific response. Antigen-specific responses can also be detected by assaying production of specific cytokines such as interleukin 2 (IL-2), IL-4, interferon-gamma, etc.

HIV-specific proliferative responses can be demonstrated in patients with acute HIV infection (70) but are quickly lost as a result of ongoing virus replication and depletion of HIV-specific CD4 cells. Proliferation in response to antigens from opportunistic pathogens such as candida, PCP, MAC, CMV, etc can also be demonstrated. These responses are lost as HIV disease progresses, and are gradually restored in patients after initiation of potent antiretroviral therapy (71; 72).

However, the correlation between pathogen-specific levels of proliferation in vitro and the risk of disease in vivo has not been established. Cells can also be stimulated to proliferate non-specifically by complex plant carbohydrates (lectins) such as phytohemagglutinin (PHA). PHA-dependent proliferation is usually preserved until late stages of HIV disease.

4/15/01

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