Accepting clocks that tell time poorly: fluid-phase versus standard ELISA autoantibody assays.
The predominant autoantibody assays employed in basic immunologic studies are variations of solid-phase assays where autoantigens are bound to 96-well plates. Though the assay format is convenient and often appropriate for studies of induced immune responses in inbred strains of mice, we will argue that this assay format usually, but not always, leads in clinical medicine to what should be unacceptable false positive results as well as lower sensitivity compared to the current generation of high throughput fluid-phase radioassays. Utilizing simple in vitro transcription and translation labeling of autoantigens, it is now possible to rapidly create fluid-phase radioassays for most (but not all) autoantigens, thereby allowing direct comparison between the different assay formats. In addition, adding a fluid-phase competition step to both solid-phase ELISA assays and even fluid-phase radioassays can enhance specificity. Development in a field of such assays with excellent specificity and sensitivity (e.g. studies of type 1A diabetes) is fostered by Societies sponsoring workshops where blinded samples are evaluated with "competing" assay formats for sensitivity, specificity, and reproducibility.
Liu
Edwin
E
Barbara Davis Center for Childhood Diabetes, University of Colorado Denver Health Sciences Center, Aurora, Colorado, USA. Edwin,Liu@uchsc.edu
Eisenbarth
George S
GS
D001323:Autoantibodies / Q000032:analysis* / Q000097:blood / Q000276:immunology; D001324:Autoantigens / Q000737:chemistry / Q000276:immunology; D001327:Autoimmune Diseases / Q000175:diagnosis / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000592:standards; D006801:Humans; D011863:Radioimmunoassay / Q000379:methods* / Q000592:standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D001323:Autoantibodies; D001324:Autoantigens