Molecular beacons can detect B. burgdorferi between 1 and 106 in multiplex assay, when C3H mouse DNA was also included. Amplification plots of recA and nidogen genes in PCR assays to estimate quantities of B. burgdorferi (A) and mouse (C) DNA are shown. Uninfected mouse heart DNA (containing 105 nidogen copies) spiked with ten-fold dilutions of B. burgdorferi strain N40 ranging from 1 to 106 were used in the PCR assays containing both RecA3 and Nidogen molecular beacons. Sensitivity and specificity of the detection system is indicated by the ability of RecA3 and Nidogen molecular beacons to quantify the amplicons from both the recA and the nidogen genes in the same PCR assay tubes. A high coefficient of correlation (r2 = 0.996) between the Ct values and the spirochete number obtained from the standard curve (B) indicates that the molecular beacons can be used effectively to quantify spirochete burden in infected tissues using multiplex assay system.