A comparison of fluorescent SSCP and denaturing HPLC for high throughput mutation scanning

We examined 67 different mutations in 16 different amplicons in a compariso n of mutation detection by fluorescent single strand conformation polymorph ism (F-SSCP) and by denaturing HPLC (DHPLC). F-SSCP was used to analyze flu orescent amplicons with internal size standards and automated fragment anal ysis (GeneScan, PE Applied Biosystems, Foster City, CA). Zn DHPLC, unlabell ed amplicons were analyzed by reverse phase HPLC with fragment detection by absorbance at 260nm. Both methods had high sensitivity (95-100%) and speci ficity (100%), Overall, F-SSCP with external temperature control was the mo re sensitive method, but DHPLC was particularly useful for the rapid analys is of novel fragments. Hum Mutat 15:556-564, 2000. (C) 2000 Wiley-Liss, Inc .