Advanced Techniques in Diagnostic Microbiology

TABLE 4.1. Types of antibody detection methods.
Sample Lable conjugated on
Method Capture antigen (Ag) target antibody detector (secondary) antibody Detection method
Colorimetric or chromogenic Usually goat, mouse, or rabbit anti-human IgG
Latex agglutination Ag on bead suspension Serum Visible agglutination
ELISA Ag on solid phase or
microparticle
Serum Enzyme (HRP or AP a ) Visible color, optic density
(OD) by spetrophotometer
Immunoblotting Ag on nitrocellulose
membrane
Lateral flow diffusion
(handheld)
Ag colloidal gold-labeled on
nitrocellulose or nylon
membrane
Serum Enzyme (HRP or AP a ) Visible band
Serum, blood,
oral fluid
Colloidal gold (chromatographic lateral flow) Visible line
Radioimmunoassay (RIA) Ag on bead (or radiolabeled) Serum radiolabeled ( 125 I, 14 C, 3 H) Radioactivity by gamma
counter
Chemiluminescence (CLIA)
Enhanced CLIA
Electrochemiluminescence
(ECL)
Fluorescence Indirect
fluorescence (IFA)
Time-resolved
fluoroescence (TRF)
Flow cytometry (FC)
Multianalyte profile
(xMAP)
Ag on solid phase or
microparticle
Serum Luminol (dioxetane through HRP or AP a )or
acridinium ester
Photon output or light by
luminometer
Ag on magnetic beads Serum Chelate ruthenium (Ru) as electron carrier (TPA as substrate)
Photon output by flow cell
with photon detector
Ag bound on slide or
Serum Fluorescein isothiocyanate (FITC) conjugated Fluorescence by microscope
microparticle
under UV light or fluoromter
Ag bound on microparticle Serum Fluorescein biotinylated with lanthanide Fluorescence by fluorometer
chelate (europium) in low pH
Ag-coated dyed microsphere Serum Fluorescein (through biotin-Ab to streptavidin) Fluorescence cell scanner
(flow cytometer), or with
FlowMetrix (Luminex)
a Horseradish peroxidase (HRP) or alkaline phosphatase (AP).
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