Microfluidic device for rapid digestion of tissues into cellular suspensions

Abstract

The ability to harvest single cells from tissues is currently a bottleneck for cell-based diagnostic technologies, and remains crucial in the fields of tissue engineering and regenerative medicine. Tissues are typically broken down using proteolytic digestion and various mechanical treatments, but success has been limited due to long processing times, low yield, and high manual labor burden. Here, we present a novel microfluidic device that utilizes precision fluid flows to improve the speed and efficiency of tissue digestion. The microfluidic channels were designed to apply hydrodynamic shear forces at discrete locations on tissue specimens up to 1 cm in length and 1 mm in diameter, thereby accelerating digestion through hydrodynamic shear forces and improved enzyme-tissue contact. We show using animal organs that our digestion device with hydro-mincing capabilities was superior to conventional scalpel mincing and digestion based on recovery of DNA and viable single cells. Thus, our microfluidic digestion device can eliminate or reduce the need to mince tissue samples with a scalpel, while reducing sample processing time and preserving cell viability. Another advantage is that downstream microfluidic operations could be integrated to enable advanced cell processing and analysis capabilities. We envision our novel device being used in research and clinical settings to promote single cell-based analysis technologies, as well as to isolate primary, progenitor, and stem cells for use in the fields of tissue engineering and regenerative medicine.

Novel microfluidic platform accelerates processing of tissue into single cells for molecular analysis and primary culture models

“…We initially tested ow rates of 10 and 20 mL/min, which were used in previous work. 52,54,55 After 15 or 60 min of recirculation, sample was collected, washed, and genomic DNA (gDNA) was extracted to assess total cell recovery. A control was minced and gDNA was directly extracted to provide an upper recovery limit.…”

“…We then developed a device for on-chip tissue digestion using the combination of shear forces and proteolytic enzymes. 54 Finally, we developed a lter device containing nylon mesh membranes that removed large tissue fragments, while also dissociating smaller cell aggregates and clusters. 55 The micro uidic digestion, dissociation, and lter devices each enhanced single cell recovery when operated independently.…”

Microfluidic Devices in the Fast‐Growing Domain of Single‐Cell Analysis

“…The conventional methods of tissue dissociation include titration, manual cutting with scalpel, micro‐scissors, tissue choppers and micro fabricated filters put the sample at high risk of contamination and require laborious work. Microfluidic devices (both enzymatic‐based and mechanical‐based devices) make this process automated and easy with high yield and viability of single‐cells . The controllable laminar flow in microfluidic devices offers high reproducibility and the closed environment reduced the risk of foreign contamination.…”

“…Reprinted with permission from Ref. [17].Copyright 2017 Royal Society of Chemistry. ( e) Schematic of FSS driven tissue dissociation.Reprinted with permission from Ref.…”

“…Microfluidic devices (both enzymatic-based and mechanical-basedd evices)m ake this process automated and easy with high yield and viability of single-cells. [17] The controllable laminarf low in microfluidic devices offersh igh reproducibility and the closed environment reduced the risk of foreign contamination. The mechanical dissociation methodi s of particulari nterestb ecause in this techniquet he isolated single-cells maintain the endogenous surface marker expression.…”