학술논문

A high‐throughput microfluidic mechanoporation platform to enable intracellular delivery of cyclic peptides in cell‐based assays
Document Type
article
Source
Bioengineering & Translational Medicine, Vol 8, Iss 5, Pp n/a-n/a (2023)
Subject
automation
cell‐based assays
cyclic peptides
intracellular delivery
microfluidics
protein–protein interactions
Chemical engineering
TP155-156
Biotechnology
TP248.13-248.65
Therapeutics. Pharmacology
RM1-950
Language
English
ISSN
2380-6761
Abstract
Abstract Cyclic peptides are poised to target historically difficult to drug intracellular protein–protein interactions, however, their general cell impermeability poses a challenge for characterizing function. Recent advances in microfluidics have enabled permeabilization of the cytoplasmic membrane by physical cell deformation (i.e., mechanoporation), resulting in intracellular delivery of impermeable macromolecules in vector‐ and electrophoretic‐free approaches. However, the number of payloads (e.g., peptides) and/or concentrations delivered via microfluidic mechanoporation is limited by having to pre‐mix cells and payloads, a manually intensive process. In this work, we show that cells are momentarily permeable (t1/2 = 1.1–2.8 min) after microfluidic vortex shedding (μVS) and that lower molecular weight macromolecules can be cytosolically delivered upon immediate exposure after cells are processed/permeabilized. To increase the ability to screen peptides, we built a system, dispensing‐microfluidic vortex shedding (DμVS), that integrates a μVS chip with inline microplate‐based dispensing. To do so, we synced an electronic pressure regulator, flow sensor, on/off dispense valve, and an x‐y motion platform in a software‐driven feedback loop. Using this system, we were able to deliver low microliter‐scale volumes of transiently mechanoporated cells to hundreds of wells on microtiter plates in just several minutes (e.g., 96‐well plate filled in