Method Description

Antinuclear Antibodies, HEp-2 Substrate:

Antibodies to nuclear antigens in a human epithelial type 2 (HEp-2) cell line by an indirect immunofluorescent technique. Commercial slides prepared from HEp-2 cells are used as a substrate. IgG antibodies in serum samples are detected after incubation of serum with the commercial slides by the addition of a fluorescein isothiocyanate (FITC)-labeled antihuman-IgG reagent. All patient samples are initially screened at 1:80.(Package insert: NOVA Lite DAPI ANA. Inova Diagnostics; 06/2018)

Scl 70 Antibodies:

Recombinant Scl-70 antigen is bound to polystyrene microspheres, which are impregnated with fluorescent dyes to create a unique fluorescent signature. Scl-70 antibodies, if present in diluted serum, bind to the Scl-70 antigen on the microspheres. The microspheres are washed to remove extraneous serum proteins. Phycoerythrin-conjugated antihuman IgG antibody is then added to detect IgG anti-Scl-70 bound to the microspheres. The microspheres are washed to remove unbound conjugate, and bound conjugate is detected by laser photometry. A primary laser reveals the fluorescent signature of each microsphere to distinguish it from microspheres that are labeled with other antigens, and a secondary laser reveals the level of PE fluorescence associated with each microsphere. Results are calculated by comparing median fluorescence response for Scl-70 microspheres to a 4-point calibration curve.(Package insert: Bioplex 2200 ANA Screen. Bio-Rad Laboratories; 02/2019)

RNA Polymerase III Antibodies:

The immunodominant fragment of RNA polymerase III antigen is derived from recombinant DNA technology. Purified RNA polymerase III antigen is adsorbed to the wells of a polystyrene microtiter plate under conditions that preserve the antigen in its antigenic state. Prediluted controls and diluted patient sera are added to separate wells. Unbound sample is washed away, and an enzyme-labeled antihuman IgG conjugate is added to each well. After incubation and washing away of unbound enzyme-labeled antihuman IgG, the bound conjugate is measured by adding a chromogenic substrate. The intensity of the absorbance produced is measured with an automated microwell plate reader. Results are calculated by comparison to a single-point calibrator.(Package insert: QUANTA Lite RNA Pol III. INOVA Diagnostics; 05/2019)