Method Description

The BioPlex 2200 Syphilis Total and RPR kit employs Treponema pallidum fusion protein (rTP47/rTP17) and cardiolipin antigen-coated fluoromagnetic beads with unique fluorescent signatures to identify the presence of IgG and IgM antibodies to T pallidum and nontreponemal reagin antibodies in a 2-step assay format. Dyed beads are coated with recombinant T pallidum rTP47/rTP17 fusion protein or cardiolipin antigen. The BioPlex 2200 System combines an aliquot of patient sample, sample diluent, and bead reagent into a reaction vessel. The mixture is incubated at 37° C. After a wash cycle, a mixture of murine monoclonal anti-human IgG and murine monoclonal anti-human IgM antibody conjugated to phycoerythrin (PE) is added to the dyed beads, and this mixture is incubated at 37° C. The excess conjugate is removed in another wash cycle, and the beads are re-suspended in wash buffer. The bead mixture then passes through the detector. The identity of the dyed beads is determined by the fluorescence of the dyes, and the amount of antibody captured by the antigen is determined by the fluorescence of the attached PE. Raw data is calculated in relative fluorescence intensity (RFI).(Package insert: BioPlex 2200 Syphilis Total and RPR. Bio-Rad; 06/2017)

If the total antibody result is reactive, a rapid plasma reagin (RPR) screen is performed on the BioPlex 2200. If the RPR screen is reactive the RPR titer is performed. The RPR titer test is a macroscopic screening assay done with unheated serum. Reagin reacts with nontreponemal antigen containing colloidal charcoal particles. This reaction results in a visual flocculation of the black particles against the white card background. The test yields a positive or negative result, and all positive samples are titered to determine the highest positive dilution.(Huber TW, Storms S, Young P, et al: Reactivity of microhemagglutination, fluorescent treponemal antibody absorption, Venereal Disease Research Laboratory, and rapid plasma reagin tests in primary syphilis. J Clin Microbiol. 1983 Mar;17[3]:405-409; Kaur G, Kaur P: Syphilis testing in blood donors: an update. Blood Transfus. 2015 Apr;13[2]:197-204)

If the RPR screen is negative, the Serodia Treponema pallidum particle agglutination (TP-PA) test is performed. The TP-PA test is based on the agglutination of colored gelatin particle carriers sensitized with T pallidum (Nichols Strain) antigen. Serum samples are serially diluted in microplate wells. Sensitized gelatin particles are added to respective wells and the contents of the plate mixed. The mixture is incubated for 2 hours at ambient temperature. Serum containing specific antibodies will react with the antigen-sensitized colored gelatin particles to form a smooth mat of agglutinated particles in the microplate well. A compact button formed by the settling of the non-agglutinated particles characterizes negative reactions. The agglutination patterns are read visually to determine interpretation.(Package insert: Serodia TP-PA. Fujirebio Diagnostics, Inc; 04/2015)