Development of a reliable low-cost controlled cooling rate instrument for the cryopreservation of hematopoietic stem cells

Abstract

An optimal cooling rate is one of the critical factors influencing the survival of cells during cryopreservation. In this paper we describe a novel device, named the box-in-box, which was developed for optimal cryopreservation of human hematopoietic stem cells (HSC). This work presents the design of the device, a mathematical formulation describing the expected temperature histories of samples during the freezing process, along with actual experimental results of thermal profile tests. In experiments, when the box-in-box device was transferred from room temperature to a −80 °C freezer, a cooling rate of −1~−3.5 °C/min, which has been widely used for the cryopreservation of HSC, was achieved. In order to further evaluate this device, HSC cryopreservation was compared between the box-in-box device and a commercially available controlled rate freezer (CryoMed). The experimental data, including total cell population and CD34+ hematopoietic progenitor cell recovery rates, viability, and cell culture colony assays, showed that box-in-box worked as well as CryoMed instrument. There was no significant difference in either survival rate or the culture/colony outcome between the two devices. In conclusion, the box-in-box device can work as a cheap, durable, reliable and maintenance-free instrument for the cryopreservation of HSC. This concept of a box-in-box may also be adapted to other cooling rates to support cryopreservation in a wide variety of tissues and cells.

Cryopreservation of Mycobacterium tuberculosis Complex Cells

“…The BIB method is inexpensive, easy to perform, and reliable and delivers a reproducible cooling rate. The BIB device has been successfully used for cryopreservation of stem cells (21,25,26). The results in this study showed that it delivers the highest recovery of M. tuberculosis after cryopreservation.…”

“…1), a thermal insulation material made of polyethylene fills an aluminum enclosure. The thickness of the polyethylene layer was designed for a low cooling rate and then demonstrated empirically (21). For cryopreservation in BIB, vials were placed in the slots cut into the polyethylene foam.…”

Sensing and Sensibility: Single‐Islet‐based Quality Control Assay of Cryopreserved Pancreatic Islets with Functionalized Hydrogel Microcapsules

“…A custom-designed freezing device, box-in-box (BIB), was applied for the slow cooling of the samples. [ 25,26 ] The microcapsules with single islets were cryopreserved and islet functionalities (adenosine triphosphate (ATP) and insulin secretion) were assessed after the freezing and thawing steps. [ 27 ] The OCR of the frozen/thawed individualislet capsules were also analyzed by monitoring the real-time oxygen uptake of each islet, based on the fl uorescent intensity of FOSD embedded in the hydrogel capsules.…”

“…The BIB system is a simple and reliable passive cooling rate-controlled device, which can provide cooling rate at about −1 °C min −1 . [ 25,26 ] After storage in the −80 °C freezer for 1 or 7 d, the BIB was taken out of the freezer and the frozen samples were thawed in the thermostatic water bath (37 °C) quickly with agitation until a very tiny ice crystal remained inside. To remove CPA after thawing, the precooled (4 °C) islet culture medium was added dropwise and slowly to the islet suspension, with the fi nal volume ratio of 10:1.…”

On-Board Array for Multiplexed Semi-Active Cooling-Rate-Controlled Cryopreservation of Living Cells

“…The optimal cooling rates of different cell types vary remarkably due to their distinct cryobiological characteristics [10]. Cell types with low optimal cooling rates (\10°C/min), such as hematopoietic stem cells [1,11], immune cells [12,13], and mammalian ova [14], require much more careful treatment than those with high optimal cooling rates ([100°C/min), such as red blood cells [15], during their freezing processes to ensure maximal survival rates.…”