Multifunctional Screening Platform for the Highly Efficient Discovery of Aptamers with High Affinity and Specificity

Abstract

Aptamers have attracted much attention as the next generation of affinity reagents. Unfortunately, the selection efficiency remains a critical bottleneck for the widespread application of aptamers. Herein, to accelerate aptamers discovery, a multifunctional microfluidic selection platform was developed, on which the selection efficiency was greatly improved and high-affinity and -specificity aptamers were generated within two round selections. The multifunctional screening platform, precisely manipulating magnetic beads on the micrometer scale, improved selection performance based on microfluidic continuous flow and enhanced the selection process control via in situ monitoring and real-time evaluation. This method could suppress ∼50-fold nonspecific binding nucleic acids compared to the conventional methods, further eliminate weakly bound nucleic acids within 9 min, and simultaneously perform the negative selection and positive selection. And the selection effectiveness was in situ and real-time monitoring. Three aptamers showed high affinity and specificity toward mucin 1 (MUC1) with dissociation constants (K) in nanomolar range (from 22 to 65 nM). Furthermore, the selected aptamer was able to specially label cancer cells and efficiently capture exosomes with 64% capture efficiency. It demonstrated that the multifunctional screening platform was an efficient method to generate high-quality aptamers in a rapid and economic manner.

Microfluidic methods for aptamer selection and characterization

“…Recently, a device by Hong et al . 79 incorporated both positive and negative selection chambers into the chip separated by a microvalve, reducing the time and hassle of performing a negative selection. A quantitative fluorescence binding analysis was performed on a separate microfluidic chip using magnetic target immobilization, demonstrating a trend towards miniaturization of additional steps of the aptamer selection process.…”

“…For example, the magnetic microbead SELEX by Hong et al . 79 was presented with a custom chip that measured the fluorescence intensity of each magnetically bound sample post-partitioning, performing a microfluidic version of the normal microplate fluorescence binding assay. There is the potential for future integration of this type of technique directly into the selection, particularly when the partitioning already makes use of magnetic beads as well.…”

Simultaneous and automated detection of influenza A virus hemagglutinin H7 and H9 based on magnetism and size mediated microfluidic chip

“…Moreover, it is difficult to simultaneously use different-size magnetic beads (from nanometers to micrometers) in biological applications due to their similar magnetic magnitudes. Microfluidic technology has many unique advantages over conventional analytical techniques, which could reduce sample and reagent consumption, high throughout, miniaturization, multiple physical field manipulation, and the potentiality of automation [22]. Thus, many efforts have been devoted to the development of magnetic-based assays on microfluidic chips [4,[23][24][25][26].…”

“…In our previous work, we have developed some biomedical applications based on magnetic beads on the microfluidic chip [4,22,[27][28][29][30][31], and also dedicating ourselves to the construction of microfluidic devices [32][33][34]. On this previous basis, a magnetism and size mediated multiphysics field microfluidic chip was constructed for simultaneous and automated detection of multiple influenza HA (Scheme 1).…”

Quantitative Determination of the Carboxyl Groups on Individual Nanoparticles by Acid‐Base Titrimetry

“…For instances, the fluorescent and magnetic nanoparticles prepared by pang's groups had carboxyl groups on their surfaces and could retain good monodispersibility for at least 12 months . On the other hand, carboxyl groups can be conveniently conjugated with various molecules, such as proteins, DNAs, and so on . Therefore, it is very important to develop a simple and rapid method for quantifying the number of carboxyl groups on individual nanoparticles, for which, two problems need to be solved.…”