Reverse-phase HPLC purifies all PentaBase probes to give the purest products. Primers are also purified by reverse-phase HPLC and desalting and removing the final DMT group. Desalting removes residual by-products from the different processes.
Every probe from PentaBase is strictly controlled for quality and fluorescence potential in an affinity test. To do this test, we synthesize a complementary oligo target and run a melting curve from 95°C to 35°C. The figure shows the result of such a test. The red curve shows the probe in the presence of its complementary target, and the yellow line shows the probe without a target. Halfway down, the dip in fluorescence of the red line corresponds to the melting temperature of the duplex. The exact melting temperature of the probe-target duplex is found by calculating the inverted derivative of the curve.