Immunology and Serology
Immunology and serology tests antibody structure and function, assay formats, and the algorithms used to interpret infectious disease serology and autoimmune testing. The exam emphasizes antibody kinetics, prozone effects, and confirmatory testing.
Immune fundamentals
IgM is the first antibody of the primary response; it is a pentamer, activates complement strongly, and does not cross the placenta. IgG dominates the secondary response and is the only immunoglobulin that crosses the placenta. IgA is mucosal and appears as secretory IgA in secretions. IgE mediates type I hypersensitivity. Agglutination and precipitation require lattice formation, which peaks at the zone of equivalence; antibody excess produces a prozone and antigen excess a postzone, both giving false-negative results.
Assay formats
ELISA formats include direct, indirect, sandwich, and competitive, all using an enzyme conjugate and substrate. Immunofluorescence uses tissue or cell-based slides, as in ANA testing. Chemiluminescence offers high sensitivity over a wide range. Nephelometry quantifies immunoglobulins and complement. Western blot is a confirmatory method. Flow cytometry enumerates CD4 cells and phenotypes leukemias. Molecular methods such as PCR detect nucleic acid directly.
Serologic algorithms
Syphilis reverse algorithm: screen with a treponemal EIA or CIA, reflex positives to RPR, and confirm discordant results with a second treponemal test such as TP-PA. HIV screening uses a fourth-generation antigen/antibody combination assay with differentiation for reactive results. Hepatitis B markers: HBsAg indicates current infection, anti-HBs indicates immunity, and anti-HBc IgM indicates acute infection. Heterophile antibody tests are often negative in young children with mononucleosis.
Worked example
An undiluted RPR is nonreactive, but the treponemal EIA is positive and the patient has a compatible history. Repeat the RPR at 1:8 and 1:16; reactivity appears only at higher dilution, demonstrating prozone from antibody excess. Report the titer and note the prozone.
Traps that cost marks
- Missing a prozone and reporting a false-negative agglutination result.
- Interpreting IgM before the window period closes.
- Passive antibody from transfusion or IVIG producing a false-positive result.
- Rheumatoid factor causing false-positive latex agglutination tests.
- Confusing ANA titer with ANA pattern, which are separate reports.