Total integrated centrifugal genetic analyzer for point-of-care Covid-19 testing with automatic and high-throughput capability

Abstract

We propose a prototype portable genetic analyzer, which performs the entire processes including RNA purification, reverse transcription multiplex loop-mediated isothermal amplification (RT-LAMP), and real-time fluorescence detection. Ten units of integrated functional entity were incorporated on a centrifugal disc, so that 10 samples could be analyzed on single device in one run. The 10 aliquoting chambers were connected sidewards by the zigzag aliquoting channel and were linked to the glass filter column outwards via the passive valves for the RNA extraction. The lower sideward capillary pressure of the zigzag aliquoting channel than the outward capillary pressure of the passive values is capable of automatic division of solution into the 10 aliquoting chambers with one injection by a needle. The sequent loading and partitioning of the sample, the washing, and the elution solutions by centrifugal protocols rendered the RT-LAMP cocktails ready for amplification in the three reaction chambers for each sample, in which the RT-LAMP primers targeting orf1ab, N and S genes were dried. After blocking the reaction chambers by wax to prevent evaporation, RT-LAMP reaction proceeded at 63 °C, and all the positive fluorescent signals in the three chambers verifies the existence of Covid-19 in the sample. For point-of-care testing (POCT), we constructed a portable workstation that consists of a solution storage, a solution injection system, a spinning motor, and a fluorescence detector. Using the proposed platform, we could perform the Covid-19 molecular diagnostics for 10 samples per run on single device in an automatic and high-throughput manner in 1.5 h.