US Patent 11530438 Highly specific circular proximity ligation assay

Provided herein is a circular proximity ligation assay in which proximity-probes are employed as bridges to connect two free oligonucleotides via a dual ligation event, resulting in the formation of a circle. The circles are then quantified by, e.g., qPCR. The addition of an extra oligonucleotide is believed to enhance specificity by decreasing the probability of random background ligation events. In addition, circle formation may have selective advantages, as uncircularized DNA can be removed by a simple exonuclease treatment and it has streamlined the workflow by eliminating preamplification prior to qPCR.