Known as RTF-EXPAR (Reverse Transcription-Free - EXPonential Amplification Reaction), it avoids the ‘reverse transcription’ method of converting to RNA to DNA, which both PCR and LAMP tests rely on. Instead, the Birmingham test uses a new RNA-to-DNA conversion step, pairing it with an alternative method for DNA amplification. Combined, the two strands of RTF-EXPAR give a sample-to-signal time of under 10 minutes, even for low viral levels. The research is published in Proceedings of the National Academy of Sciences.
“Both the reverse transcription and amplification steps slow down existing COVID assays that are based on nucleic acid detection, compared to antigen tests, such as lateral flow, which do not have these steps,” explained Professor Tim Dafforn, from Birmingham’s School of Biosciences.
However, while this makes lateral flow tests faster than those based on PCR and LAMP, in return they are typically less sensitive. An ideal test would be one that is both sufficiently sensitive and speedy – our test, called RTF-EXPAR, achieves this goal.”
The study revealed that the RTF-EXPAR method converts under 10 strands of RNA into billions of copies of DNA in under 10 minutes, using a one-pot assay that is compatible with more basic, benchtop equipment than that used with current testing methods.
RTF-EXPAR also demonstrated significant improvements over both PCR and LAMP-based assays on time to signal detection. At low concentrations of RNA (7.25 copies/µL), the time to signal detection was 42.67 (± 0.47) minutes for PCR, 11.25 (± 0.20) minutes for LAMP, and 8.75 (± 0.35) minutes for EXPAR. At high (1450 copies/µL) concentrations of viral RNA, the time to signal detection was 34.00 (± 0.00) minutes for PCR, 11.25 (± 0.20) minutes for LAMP, and 3.08 (± 0.42) minutes for EXPAR.
“EXPAR amplifies DNA at a single temperature, thus avoiding lengthy heating and cooling steps found in PCR,” said Professor Dafforn.
“However, while LAMP also uses a single temperature for amplification, EXPAR is a simpler and a more direct process, in which much smaller strands are amplified. This makes EXPAR an even faster DNA amplification technique than not only PCR but also LAMP.”
The Birmingham team is currently seeking partners to help commercialise the technology and roll it out as widely as possible.