Method Description

Recombinant SS-A/Ro 52 kDa, affinity-purified SS-A/Ro 60 kDa, and affinity-purified SS-B antigen are coupled covalently to polystyrene microspheres that 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, and SS-B/La antibodies, if present, 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-A/Ro or 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-A/Ro and SS-B/La microspheres to a 4-point calibration curve.(Package insert: Bioplex 2200 ANA Screen. Bio-Rad Laboratories; 02/2019)