Search

MicroRNAs (miRNAs) are endogenous molecules with regulatory functions. The purification and enrichment of miRNA are essential for its precise and sensitive detection. miRNA isolated using commercial kits contains abundant interfering RNAs, and the concentration of miRNA may not be adequate for detection. Herein, we prepared a reduced graphene oxide (rGO)-based magnetic solid-phase extraction material for the enrichment and ultrasensitive detection of miRNA from intricate nucleic acid solutions. In situ reverse transcription (RT) was developed as the most efficient approach to desorb miRNA from rGO among the methods that are compatible for the subsequent amplification reported thus far. Additionally, rolling circle amplification and qPCR were used to detect let-7a with a decrease of the limit of detection by 24.7- and 31.3-fold, respectively. This material was also successfully used to extract and detect miRNA from total RNA isolated from human plasma. Our results show that the material prepared in this study has the potential for cancer biopsy in clinics and the discovery of new miRNAs in scientific research.

Antibiotic residues and illegal additives are among the most common contaminants in milk and other dairy products, and they have become essential public health concerns. To ensure the safety of milk, rapid and convenient screening methods are highly desired. Here, we integrated microarray technology into a microfluidic device to achieve rapid, sensitive, and fully automated detection of chloramphenicol, tetracyclines, enrofloxacin, cephalexin, sulfonamides, and melamine in milk on a centrifugal microfluidic platform with two rotation axes. All the liquid reagent for the immunoassay was prestored in the reagent chambers of the microdevice and can be released on demand. The whole detection can be automatically accomplished within 17 min, and the limits of detection were defined as 0.92, 1.01, 1.83, 1.14, 1.96, and 7.80 μg/kg for chloramphenicol, tetracycline (a typical drug of tetracyclines), enrofloxacin, cephalexin, sulfadiazine (a typical drug of sulfonamides), and melamine, respectively, satisfying the national standards for maximum residue limits in China. Raw milk samples were used to test the performance of the current immunoassay system, and the recovery rates in the repeatability tests ranged from 80 to 111%, showing a good performance. In summary, the immunoassay system established in this study can simultaneously detect six contaminants of four samples in a fully automated, cost-effective, and easy-to-use manner and thus has great promise as a screening tool for food safety testing.

The genotyping of SNPs (single nucleotide polymorphisms) is a prerequisite for the analysis of many genetic diseases, including hereditary hearing-loss. However, the existing methods for SNP detection suffer from a long detection period, tedious operation, and a high risk of carryover contamination. To address these challenges, a microfluidic chip is constructed for rapid and efficient SNP genotyping by dividing the sample into many independent chambers for Kompetitive Allele Specific PCR in this study. Using this strategy, multiple detection can be easily accomplished and the challenge for the establishment of multiplex PCR is fundamentally overcome. The entire detection can be finished within 2 h in a fully sealed manner with this method, which is quite simple compared to SNaPshot and MassArray. After assessment of the basic performance, this chip was applied to screen 15 mutations, including SNPs and InDels (insertion-deletion markers), that can cover more than 80% of cases of hereditary hearing-loss in China. Over 40 clinical samples were analyzed with this microfluidic chip for SNP genotyping, and the results are consistent with that obtained by Sanger sequencing, demonstrating its practicability and potential in the application of genetic disease detection.

Lateral flow immunoassay (LFIA) is widely used but is limited by its sensitivity. In this study, a novel centrifugation-assisted lateral flow immunoassay (CLFIA) was proposed that had enhanced sensitivity compared to traditional LFIA based on test strips. For CLFIA, a vaulted piece of nitrocellulose membrane was prepared and inserted into a centrifugal disc. Powered by the centrifugal force, the sample volume on the disc was not limited and the flow rate of the reaction fluid was steady and adjustable at different rotation speeds. It was found that lower rotation speeds and larger sample volumes resulted in greater signal intensity in the nitrocellulose membrane as well as higher sensitivity, indicating that the actively controlled flow on the disc allowed for sensitivity enhancement of CLFIA. To operate CLFIA on the centrifugal disc, a portable and cost-effective operating device was constructed to rotate the disc with a stepper motor and collect the results with a smartphone. The proposed method was successfully applied to detect prostate specific antigen (PSA) in human serum. Standard curves were established for CLFIA and LFIA, and both had correlation coefficients of up to 0.99. Under optimal conditions (1500 rpm rotation speed, 120 μL sample volume), the detection limit of CLFIA reached 0.067 ng/mL, showing a 6.2-fold improvement in sensitivity compared to that of LFIA. With clinical serum samples, a good correlation was observed between PSA concentrations measured by CLFIA and by a bulky commercial instrument in hospital. In summary, this portable, cost-effective, and easy-to-use system holds great promise for biomarker detection with enhanced sensitivity compared to traditional LFIA.

Centrifugal microfluidics has been widely applied in the sample-in-answer-out systems for the analyses of nucleic acids, proteins, and small molecules. However, the inherent characteristic of unidirectional fluid propulsion limits the flexibility of these fluidic chips. Providing an extra degree of freedom to allow the unconstrained and reversible pumping of liquid is an effective strategy to address this limitation. In this study, a wirelessly charged centrifugal microfluidic platform with two rotation axes has been constructed and the flow control strategy in such platform with two degrees of freedom was comprehensively studied for the first time. Inductively coupled coils are installed on the platform to achieve wireless power transfer to the spinning stage. A micro servo motor is mounted on both sides of the stage to alter the orientation of the device around a secondary rotation axis on demand during stage rotation. The basic liquid operations on this platform, including directional transport of liquid, valving, metering, and mixing, are comprehensively studied and realized. Finally, a chip for the simultaneous determination of hexavalent chromium [Cr(VI)] and methanal in water samples is designed and tested based on the strategy presented in this paper, demonstrating the potential use of this platform for on-site environmental monitoring, food safety testing, and other life science applications.