Indirect fluorescent antibody determination (IFA) is a widely applied method of detecting diverse antibodies. For these types of tests, the antigen against which the patient makes antibody (e.g., whole Toxoplasma organisms or virus-infected tissue culture cells) is affixed to the surface of a microscope slide. The patient’s serum is diluted and placed on the slide, covering the area in which antigen was placed. If present in the serum, antibody binds to the specific antigen. Unbound antibody is then removed by washing the slide. In the second stage of the procedure, a conjugate of antihuman globulin directed specifically against IgG or IgM and a fluorescent dye (e.g., fluorescein) is placed on the slide. This labeled marker for human antibody binds to the antibody already bound to the antigen on the slide and serves as a detector, indicating binding of the antibody to the antigen when viewed under a fluorescence microscope (Figure 1). Commercially available test kits include slides coated with the antigen, positive and negative control sera, diluent for the patients’ sera, and the properly diluted conjugate. As with other commercial products, IFA systems should be used as units, without modification of the manufacturer’s instructions. Commercially available IFA tests include those for antibodies to Legionella species, B. burgdorferi, T. gondii, varicella zoster virus, cytomegalovirus, Epstein-Barr virus capsid antigen, early antigen and nuclear antigen, herpes simplex viruses types 1 and 2, rubella virus, M. pneumoniae, T. pallidum (the fluorescent treponemal antibody absorption test [FTA-ABS]), and several rickettsiae. Most of these tests, if performed properly, give extremely specific and sensitive results. Proper interpretation of IFA tests requires experienced and technically competent technologists. These tests can be performed rapidly and are cost effective.

Fig1. Indirect fluorescent antibody tests for Toxoplasma gondii, IgG antibodies. A, Positive reaction. B, Negative reaction. (Courtesy  Meridian, Cincinnati, Ohio.)

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