Using probes labeled with different reporter dyes allows the simultaneous detection and quantification of multiple target genes in a single (multiplex) reaction.
Multiplex qPCR can be advantageous when the available template is limited, by maximizing the number of amplifications that can be performed. Further, when there are large numbers of samples, multiplexing can help save costs by reducing the total number of reactions required. However, developing a multiplex assay can bring new challenges, such as more complex design requirements. It can be difficult to design fully compatible probe and primer sets, and this is exacerbated as the number of products to be detected is increased. In addition, optimizing the multiplex qPCR conditions to suit all primer/probe sets can lead to a reduction in sensitivity of detection, as all primer/probe sets need to have similar reaction kinetics and buffer requirements. For assistance in designing or trouble-shooting your qPCR probes, please contact us through our complimentary probe design service.