Last Updated on November 28, 2023

Freezing of cells
Cryopreservation is a crucial technique in cell research, allowing us to store valuable cells for extended periods while preserving their viability and functionality. However, without a proper freeze protocol, the freezing process can be damaging to cells, resulting in reduced cell viability and compromised research outcomes. To ensure the best results, it is essential to follow a consistent freeze protocol. In this article, we will provide you with a general guideline for cryopreservation that can be tailored to different cell types. Remember to refer to specific instructions from your cell culture supplier for accurate details.

Preparing for Cryopreservation:

Before we dive into the freeze protocol, it is important to take some necessary precautions to protect both yourself and the cells. Always wear personal protective equipment and perform all procedures using aseptic techniques. Ensure that all equipment and lab materials coming into contact with the cells are properly sterilized. Working inside a laminar flow hood creates a controlled environment that minimizes the risk of contamination.

Media Exchange:

The first step in the freeze protocol involves preparing the cells for cryopreservation. Start by centrifuging the cells to collect them at the bottom of the culture vessel. Then, carefully remove the spent media, wash the cells with an appropriate buffer, and resuspend them in a cryogenic vial filled with a suitable freeze media. This freeze media should contain a cryoprotective agent such as dimethyl sulfoxide (DMSO) or glycerol. It’s worth noting that the choice of freeze media may vary depending on the cell type.

The Freezing Process:

The freezing process itself plays a critical role in cell survival and recovery. The rate at which cells cool down can significantly impact their integrity. Rapid temperature drops can lead to intracellular ice formation, which can damage the cell membrane. On the other hand, slow cooling rates may result in chemical toxicity due to higher salt concentrations.

The optimal cooling rate typically ranges around -1°C per minute. However, it is important to consider various factors such as membrane permeability and surface-to-volume ratio, as they can influence the ideal rate for different cell types. To achieve consistent results, it is advisable to use a controlled rate freezing apparatus, ensuring a gradual decrease in temperature. While some controlled rate freezers rely on liquid nitrogen, which can pose challenges and risks of mycoplasma contamination, there are alternative options available. The Strex CytoSAVER Controlled Rate Freezer, for example, offers a liquid nitrogen-free solution that simplifies the freezing process.

Alternatively, if a controlled rate freezer is not accessible, an isopropanol chamber or insulating foam can be utilized. Placing the cryovials containing cells in such a setup and leaving them overnight in a -80°C freezer attempts to mimic the recommended -1°C per minute cooling rate by slowing down heat transfer. However, it is important to note that this method often yields inconsistent results.


Once the cells are frozen, they should be promptly transferred to a long-term storage facility, such as a -80°C freezer or liquid nitrogen tank. During the transfer, it is advisable to use dry ice to maintain the cells’ frozen state. It is important to handle the cryovials carefully, as any thawing could adversely affect the cells. When ready to use the cells, they can be rapidly thawed at room temperature.

Freezing Cells Protocol – Conclusion

Following an optimal freeze protocol is crucial for the successful cryopreservation of cultured cells. By adhering to the guidelines discussed in this article and consulting specific instructions from your cell culture supplier, you can enhance cell viability and preserve their functionality for future research. If you require further information on the Strex CytoSAVER Controlled Rate Freezer or have any additional questions, please feel free to reach out to us. Our team is available to provide you with more details and assist you in finding the best cryopreservation solution for your specific needs.

Remember, by implementing a consistent freeze protocol and utilizing advanced freezing technologies like the Strex CytoSAVER Controlled Rate Freezer, you can ensure the highest quality of frozen cells, maximizing the potential of your research and preserving your valuable cell cultures for future experiments.

At Strex, we understand the importance of reliable cryopreservation techniques, and we are committed to providing innovative solutions that meet the needs of researchers like you. Contact us today and let us help you optimize your freeze protocol for the cryopreservation of cultured cells.

For more information or to request an online demo on Strex’s CytoSAVER contact us below.