Miniature auto‐perfusion bioreactor system with spiral microfluidic cell retention device

Abstract

Medium perfusion is critical in maintaining high cell concentration in cultures. The conventional membrane filtration method for medium exchange has been challenged by the fouling and clogging of the membrane filters in long‐term cultures. In this study, we present a miniature auto‐perfusion system that can be operated inside a common‐size laboratory incubator. The system is equipped with a spiral microfluidic chip for cell retention to replace conventional membrane filters, which fundamentally overcomes the clogging and fouling problem. We showed that the system supported continuous perfusion culture of Chinese hamster ovary (CHO) cells in suspension up to 14 days without cell retention chip replacement. Compared to daily manual medium change, 25% higher CHO cell concentration can be maintained at an average auto‐perfusion rate of 196 ml/day in spinner flask at 70 ml working volume (2.8 VVD). The auto‐perfusion system also resulted in better cell quality at high concentrations, in terms of higher viability, more uniform and regular morphology, and fewer aggregates. We also demonstrated the potential application of the system for culturing mesenchymal stem cells on microcarriers. This miniature auto‐perfusion system provides an excellent solution to maintain cell‐favorable conditions and high cell concentration in small‐scale cultures for research and clinical uses.

Shake tube perfusion cell cultures are suitable tools for the prediction of limiting substrate, CSPR, bleeding strategy, growth and productivity behavior

“…Thus, only a limited number of small‐scale/miniature perfusion systems have been developed, including the Ambr™15/250 (The Automation Partnership [Cambridge] Limited), 2‐6 and rocking motion (WAVE™ bioreactor from Cytiva) 7 bioreactors or the DASbox™ (Eppendorf AG) combined with ATF modules 8 . Deep‐well plates, shake tubes, Erlenmeyer and spinner flasks have also been used as perfusion models at small scale in the past 9‐15 . During perfusion medium development, SDMs are used as small surrogate cultures to estimate the ‘depth’ 16 of the medium (indicated by a low minimally supported CSPR), and to estimate the stability, metabolite, growth, and productivity behavior of the cell line, as factors influencing the product quality 13 …”

“…8 Deep-well plates, shake tubes, Erlenmeyer and spinner flasks have also been used as perfusion models at small scale in the past. [9][10][11][12][13][14][15] During perfusion medium development, SDMs are used as small surrogate cultures to estimate the 'depth' 16 of the medium (indicated by a low minimally supported CSPR), and to estimate the stability, metabolite, growth, and productivity behavior of the cell line, as factors influencing the product quality. 13 In this study, simple non-instrumented shake tubes based on a semi-continuous centrifugation perfusion protocol were investigated for their capabilities and limitations to simulate industrial perfusion processes.…”

Separation of Ultra‐High‐Density Cell Suspension via Elasto‐Inertial Microfluidics

“…Compared with other membrane‐less cell manipulation techniques such as acoustic wave separators, centrifuges, hydrocyclones, and gravity settlers, [ 51,52 ] (Table S1, Supporting Information), the microfluidic cell retention device based on elasto‐inertial cell focusing has many benefits. With its enhanced cell density capacity demonstrated in this study, the membrane‐less microfluidic device can perform cell retention with advantages including removal of small dead cells/debris and low capital/operational expenses, [ 49,50,53–55 ] which could lead to efficient and reliable high‐density perfusion culture at various scales.…”

Multi-scale model of clogging in microfluidic devices with grid-like geometries

“…For instance, in microfluidic cell sorters that sort sickle red blood cells based on their deformability, clogging leads to greater unpredictability in the device behaviour. Previous studies have shown that clogging contributes to a faster fouling of the device [6][7][8][9][10]. Investigating the detailed nature of clogging in these microfluidic devices provides an opportunity to gain a deeper understanding of their properties and how their performance can be improved.…”