Interleukin 16 (IL‐16) is a chemoattractant immunomodulatory cytokine that initiates its cellular responses through interaction with membrane‐expressed CD4. The protein may be detected by a number of methods; the choice of protocol will depend on the ultimate object of a particular experiment. The first method presented is the use of ELISA to measure IL‐16 in cell culture supernatants or biological fluids. For some applications, such as identification of IL‐16 in an unknown fluid or medium or direct assessment of its bioactivity, functional assays of IL‐16‐induced responses may be more appropriate. The chemotactic effects of IL‐16 on CD4⁺ T cells and its specific inhibition may be measured using anti‐IL‐16 antibodies; the same approach may also be applied to monocytes or eosinophils. Another effect of IL‐16 is the induction of CD25, which can be assayed using immunological staining. Finally, cell cycle progression in target cells can be measured by the incorporation of radiolabeled thymidine and confirmed by inhibition with neutralizing antibody.