Microfluidic channel optimization to improve hydrodynamic dissociation of cell aggregates and tissue

Xiaolong Qiu,Jered B Haun,Katrina M Henrikson,Jeremy A Lombardo,Pedram P Pourfard,Pulak Nath,Trisha M Westerhof,Marissa Pennell,Edward L Nelson,Jen-Huang Huang

doi:10.1038/s41598-018-20931-y

Abstract

Maximizing the speed and efficiency at which single cells can be liberated from tissues would dramatically advance cell-based diagnostics and therapies. Conventional methods involve numerous manual processing steps and long enzymatic digestion times, yet are still inefficient. In previous work, we developed a microfluidic device with a network of branching channels to improve the dissociation of cell aggregates into single cells. However, this device was not tested on tissue specimens, and further development was limited by high cost and low feature resolution. In this work, we utilized a single layer, laser micro-machined polyimide film as a rapid prototyping tool to optimize the design of our microfluidic channels to maximize dissociation efficiency. This resulted in a new design with smaller dimensions and a shark fin geometry, which increased recovery of single cells from cancer cell aggregates. We then tested device performance on mouse kidney tissue, and found that optimal results were obtained using two microfluidic devices in series, the larger original design followed by the new shark fin design as a final polishing step. We envision our microfluidic dissociation devices being used in research and clinical settings to generate single cells from various tissue specimens for diagnostic and therapeutic applications.

Highlights

Recent insights into the importance of cellular heterogeneity and rare driver cells have combined with advancements in sequencing and molecular detection technologies to help usher in the era of single cell diagnostics[1,2,3,4,5]
Extensive testing with cancer cell aggregates and spheroids demonstrated that our microfluidic device significantly improved cell recovery in terms of single cell numbers and purity
Our original microfluidic dissociation device design consisted of a network of branching channels that progressively decreased in channel width, and channels were arranged across three layers of hard PET plastic that were separated by via layers[26]

Summary

Introduction

Recent insights into the importance of cellular heterogeneity and rare driver cells have combined with advancements in sequencing and molecular detection technologies to help usher in the era of single cell diagnostics[1,2,3,4,5]. Microfluidic device technologies have been developed to aid dissociation, with early works focused on digesting, cutting, or physically breaking down tissues or cellular aggregates[23,24,25] These devices either were not designed to produce single cells, or suffered from significant clogging issues. Extensive testing with cancer cell aggregates and spheroids demonstrated that our microfluidic device significantly improved cell recovery in terms of single cell numbers and purity These results were obtained using minimal proteolytic digestion, and in some cases even without the use of enzymes. The best results are obtained using two microfluidic devices in series, with the original multilayer device first reducing large tissue fragments and aggregates into smaller units, followed by a final sample polishing step using the new optimized single layer shark fin device to produce more single cells. We will examine performance using different tissue types including various normal organs and solid tumor specimens

Methods

Results

Conclusion