Method Description

Double-Stranded DNA (dsDNA) IgG Antibodies

The test kit contains 12 microtiter strips each with 8 break-off reagent wells coated with double-stranded DNA (dsDNA). In the first reaction step, diluted patient samples, calibrators and controls are incubated in the wells. Anti-dsDNA antibodies will bind to the antigens coated in the microtiter wells. The wells are washed to remove any unbound proteins and non-specific antibodies. In a second reaction step, rabbit anti-human IgG-HRP enzyme conjugate is added to each well. The enzyme conjugate will bind to any wells that have human IgG binding to the dsDNA antigen. The wells are washed to remove any unbound HRP enzyme conjugate. 3,3,5,5, tetramethylbenzidine (TMB) enzyme substrate is added. If the HRP enzyme is present in the well (positive reaction), the HRP enzyme will react with the TMB substrate and produce a blue color. After an additional incubation time to allow the color development, a stop solution is added which turns the blue color yellow and inhibits further color development to allow for stable spectrophotometric reading. The test strips are placed in a microplate reader and the optical density of the color is measured. The amount of antigen specific bound antibody is proportional to the color intensity.(Package insert: Anti-dsDNA-NcX ELISA (IgG). EUROIMMUN; 7/8/2020)

RNP IgG Antibodies

Recombinant ribonucleoprotein particle (RNP)-68 and RNP-A antigens are bound to polystyrene microspheres, which are impregnated with fluorescent dyes to create a unique fluorescent signature. RNP antibodies, if present in diluted serum, bind to the RNP antigens on the microspheres. The microspheres are washed to remove extraneous serum proteins. Phycoerythrin (PE)-conjugated antihuman IgG antibody is then added to detect IgG anti-RNP 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 the median fluorescence response for RNP microspheres to a 4-point calibration curve.(Package insert: Bioplex 2200 ANA Screen. Bio-Rad Laboratories; 02/2019)

Scl 70 IgG 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)

Smith (Sm) IgG Antibodies

Affinity-purified Smith (Sm) antigens are bound to polystyrene microspheres, which are impregnated with fluorescent dyes to create a unique fluorescent signature. Sm antibodies, if present in diluted serum, bind to the Sm antigens on the microspheres. The microspheres are washed to remove extraneous serum proteins. Phycoerythrin (PE)-conjugated antihuman IgG antibody is then added to detect IgG anti-Sm 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 the median fluorescence response for Sm microspheres to a 4-point calibration curve.(Package insert: Bioplex 2200 ANA Screen. Bio-Rad Laboratories; 02/2019)

SS-A/Ro IgG Antibodies

Recombinant SS-A/Ro 52 kD and affinity-purified SS-A/Ro 60 kD antigens are coupled covalently to polystyrene microspheres, which are impregnated with fluorescent dyes to create a unique fluorescent signature. SS-A/Ro antibodies, if present in diluted serum, bind to the SS-A/Ro antigens on the microspheres. The microspheres are washed to remove extraneous serum proteins. Phycoerythrin (PE)-conjugated antihuman IgG antibody is then added to detect IgG anti-SS-A/Ro 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 the median fluorescence response for SS-A/Ro microspheres to a 4-point calibration curve.(Package insert: Bioplex 2200 ANA Screen. Bio-Rad Laboratories, 02/2019)

SS-B/La IgG Antibodies

Affinity-purified SS-B antigen is coupled covalently to polystyrene microspheres, which are impregnated with fluorescent dyes to create a unique fluorescent signature. SS-B/La antibodies, if present in diluted serum, bind to the SS-B antigen on the microspheres. The microspheres are washed to remove extraneous serum proteins. Phycoerythrin (PE)-conjugated antihuman IgG antibody is then added to detect IgG anti-SS-B/La 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 the median fluorescence response for SS-B/La microspheres to a 4-point calibration curve.(Package insert: Bioplex 2200 ANA Screen. Bio-Rad Laboratories; 02/2019)

Technical Interpretation

An interpretation based on the test results is generated by the laboratory information system.