Taking it to the extreme: PCR at wittwerspeed.

Abstract

If the start times of PCR thermal cycling were plotted relative to frequency, we could expect to easily see sharp peaks of start times immediately before morning/afternoon coffee breaks, lunch times, and end of day, by operators safe in the knowledge that the amplification would take 1–2 hours to complete. Perhaps it is a little faster in many, but not most, diagnostic laboratories. Indeed, careful experimental planning when I was a student allowed leisure activities at opportune times “while the PCR was running.” At the time, these PCRs were typically 50–100 μL in volume and used relatively thick 0.5-mL tubes. In the late 1990s, the Guinness Book of World Records held an entry for “fastest DNA fingerprint” of approximately 11 min (essentially rapid real-time PCR detection). Further demonstrating that faster amplification concepts are not new, the term “rapid-cycle PCR” was coined in the early 1990s to define a thermal cycling program performed in 10–30 min (1). The Guinness entry and rapid-cycle PCR have a common denominator: the world record was held by the original capillary-based LightCycler, and similar capillaries were used for the rapid-cycle PCR—both in the hands of Carl Wittwer from the University of Utah. The work of Professor Wittwer and his team is no stranger to these pages, as they previously published real-time PCR applications on the LightCycler (2) as well as early papers on the now well-adopted genotyping and variant scanning methodology of high-resolution melting analysis (HRMA) developed in his laboratory (3, 4)—work that more than justified Wittwer as the subject of an “Inquiring Minds” article, also in Clinical Chemistry (5). Since …