725Figure 31.3.Multiplex PCR and the use of fluorescently labeled microspheres to detect pathogens. A.DNA is isolat-
ed from the food sample and all prokaryotic 23S rRNA gene sequences are amplified using universal primers, which
also add a biotin label to the amplified DNA. Fluorescently labeled beads containing a different spectral pattern for
each pathogen-specific probe are added to the amplified PCR mix, and the DNA is denatured to create single-strand-
ed fragments. B.The amplified pathogenic DNA fragments bind to the beads, displaying their complementary probes.
C.The bound microspheres are labeled with fluorescent label by adding streptavidin-R-phycoerythrin, which binds to
the biotin label on the amplified DNA and fluoresces at a different wavelength than the microspheres. D.The beads
bound to the pathogenic DNA are sorted based on their spectral patterns, and the amount of fluorescence is meas-
ured to quantify the amount of each pathogen present. B.biotin; P,phycoerythrin.