C-2022: Monitoring of the residual cryoprotectant concentration with electrical conductivity measurements during cryoprotectant removal

Abstract

Cryoprotective agents (CPA) should be added to cell suspension before freezing and removed after thawing. Nowadays people mainly apply centrifugation for CPA removal. This method may cause many problems. Meanwhile, a fast, easy and cheap method to monitor the residual CPA concentration remains an unfilled need. In this work, real-time monitoring of residual CPA concentration with electrical conductivity (EC) measurements was proposed and tested. It was found that CPA concentration is correlated with the EC of the solution. The standard data of “CPA concentration—EC of the solution” for a few widely used CPA solutions were obtained. A “dilution–filtration” system based on a hollow fiber dialyzer, three peristaltic pumps and some well-designed tubing connections was applied to remove CPA. Me2SO solutions (Me2SO–NaCl–H2O solutions with or without 5% BSA) were selected as representative examples. The results showed that CPA can be removed effectively by the “dilution–filtration” system. The ECs of waste solution (filtration product) and “cell suspension” were almost identical after a short period of processing, which were validated by osmolality measurements with an osmometer. EC readings of waste solution can convey the instant residual CPA concentrations in cell suspension. This validates the feasibility of a safe (no contamination), easy and cheap way to real-time monitoring of CPA concentration in cell suspension by measuring the EC of the waste solution. Compared to other CPA removal methods that are applied traditionally (centrifugation) or proposed recently (microfluidic device), the “dilution–filtration” method may offer advantages of low time and labor consumption, low injury to the cells, high effectiveness, ease of controlling the final suspension volume and low risk of contamination. The configuration of the “dilution–filtration” system can also be modified easily to achieve CPA addition and volume control of cell suspension (concentration or dilution of the cell suspension).