Continuous microfluidic assortment of interactive ligands (CMAIL)

Abstract

Finding an interactive ligand-receptor pair is crucial to many applications, including the development of monoclonal antibodies. Biopanning, a commonly used technique for affinity screening, involves a series of washing steps and is lengthy and tedious. Here we present an approach termed continuous microfluidic assortment of interactive ligands, or CMAIL, for the screening and sorting of antigen-binding single-chain variable antibody fragments (scFv) displayed on bacteriophages (phages). Phages carrying native negative charges on their coat proteins were electrophoresed through a hydrogel matrix functionalized with target antigens under two alternating orthogonal electric fields. During the weak horizontal electric field phase, phages were differentially swept laterally depending on their affinity for the antigen, and all phages were electrophoresed down to be collected during the strong vertical electric field phase. Phages of different affinity were spatially separated, allowing the continuous operation. More than 105 CFU (colony forming unit) antigen-interacting phages were isolated with ~100% specificity from a phage library containing 3 × 109 individual members within 40 minutes of sorting using CMAIL. CMAIL is rapid, sensitive, specific, and does not employ washing, elution or magnetic beads. In conclusion, we have developed an efficient and cost-effective method for isolating and sorting affinity reagents involving phage display.

Application of microfluidic technology in antibody screening

“…Efficient enrichment and automated selection. [80,83,85] Limitation concerning the immune system. Strict control of the biopanning process.…”

“…Hsiao et al proposed a continuous microfluidic driven by two alternating orthogonal electric field drives for the screening and sorting of scFv phages without washing, elution, or magnetic bead biopanning. [85] The phage was electrophoresed on an agarose gel immobilized with the target antigen. Phage clones were enriched at different outlets mainly based on the difference in affin-ity (Figure 8D).…”

Application of Microfluidic Technology in Field of Antibody Preparation

“…Automation of microfluidic screening will speed up the phage antibody library screening process and improve efficiency, but the washing process results in the loss or inability to be enriched of clones. As shown in Figure 9D, Hsiao (Hsiao et al, 2016) proposed a continuous microfluidic driven by two alternating orthogonal electric field drives for the screening and sorting of scFv displayed on phages without washing, elution or magnetic beads. The phage was electrophoresed on an agarose gel immobilized with the target antigen, and phage clones were enriched at different outlets according to the difference in electric field force and affinity.…”

Gold Binding Peptide Identified from Microfluidic Biopanning: An Experimental and Molecular Dynamics Study

“…For the precise regulation of experimental conditions, microfluidic systems have been adopted to control the washing stringency during the phage selection. − Microfluidic biopanning systems not only require a small sample volume but also can carry out multiple steps in parallel or series in a single device. Because of their advantages, microfluidic biopanning systems have received increasing attention for screening peptides that exhibit a high binding affinity against target materials.…”