A Lab-on-a-Chip Device Integrated DNA Extraction and Solid Phase PCR Array for the Genotyping of High-Risk HPV in Clinical Samples

Abstract

Point-of-care (POC) molecular diagnostics play a crucial role in the prevention and treatment of infectious diseases. It is necessary to develop portable, easy-to-use, inexpensive and rapid molecular diagnostic tools. In this study, we proposed a lab-on-a-chip device that integrated DNA extraction, solid-phase PCR and genotyping detection. The ingenious design of the pneumatic microvalves enabled the fluid mixing and reagent storage to be organically combined, significantly reducing the size of the chip. The solid oligonucleotide array incorporated into the chip allowed the spatial separation of the primers and minimized undesirable interactions in multiplex amplification. As a proof-of-concept for POC molecular diagnostics on the device, five genotypes of high-risk human papillomavirus (HPV) (HPV16/HPV18/HPV31/HPV33/HPV58) were examined. Positive quality control samples and HPV patient cervical swab specimens were analyzed on the integrated microdevice. The platform was capable of detection approximately 50 copies of HPV virus per reaction during a single step, including DNA extraction, solid-phase PCR and genotype detection, in 1 h from samples being added to the chip. This simple and inexpensive microdevice provided great utility for the screening and monitoring of HPV genotypes. The sample-to-result platform will pave the way for wider application of POC molecular testing in the fields of clinical diagnostics, food safety, and environmental monitoring.

Point-of-care diagnostics for infectious diseases: From methods to devices

“…The device was characterized by the chaotic fluid mixing and the automatic liquid handling system, which was capable of detecting Hendra virus antibodies within 15 min and the detection limit was as low as 0.48 ng mL −1 . In another example, Zhu et al designed a LOC device that integrated DNA extraction, solid-phase PCR and genotyping detection for high-risk HPV [357] . In this device, the chip size was significantly reduced by applying the pneumatic microvalves for organically combination of the fluid mixing and reagent storage.…”

Advances in point-of-care nucleic acid extraction technologies for rapid diagnosis of human and plant diseases

“…After mechanical cell lysis, magnetic nanowires were added in the cell lysate to bind the released nucleic acids with the cationic polymer. Zhu et al (2019) designed an integrated microfluidic chip for HPV detection from cervical specimens within 1 h. The microfluidic device combined thermal lysis method of nucleic acid extraction and solid-phase PCR for rapid target detection.…”

A Novel Microfluidic Device Integrated with Chitosan-Modified Capillaries for Rapid ZIKV Detection

“…Firstly, yeast RNA medium was introduced into 11 batches of chitosan-modified capillaries, 8 cm in length. The solution in part of the capillaries was recovered during a range of time (5,10,20,30,45,60,90,120,180,240, and 300 s), and residual RNA samples were measured by a spectrophotometer (Biophotometer Plus, Eppendorf, Hamburg, Germany). The quality of adsorbed RNA was calculated by the difference between the initial concentration of the RNA and the residual one.…”

“…Reverse-transcription polymerase chain reaction (RT-PCR) is considered as a star standard for ZIKV diagnose according to the CDC [8]. However, conventional PCR progress is usually time-consuming, and requires well-trained experimenters, bulky clean bench, or commercial instruments for nucleic acids extraction, which are not suitable for filed deployment [9,10]. Point of care (POC) devices defined as instruments that can perform an analytical or diagnostic test near the site of resource-limited areas are urged to apply for epidemic disease control [11].…”