# Effect of low-frequency assisted ultrasonic on cryopreservation of L-02 hepatocyte cells

## TL;DR

This study shows that using low-frequency ultrasound during freezing improves the survival and function of liver cells, making it a promising method for cell preservation.

## Contribution

A novel ultrasonic ice-seeding system is introduced to enhance hepatocyte cryopreservation and reduce supercooling risks.

## Key findings

- Ultrasonic intensities between 0.0329 W/cm2 and 0.4316 W/cm2 achieved over 90% hepatocyte survival.
- Ultrasonic ice seeding had minimal impact on hepatocyte function over 7 days.
- The technology preserved cell secretion of urea, albumin, and glucose effectively.

## Abstract

Developing bioartificial liver and hepatocyte transplantation technology causes increasing hepatocyte cell demand. Effective long-term hepatocyte cell preservation methods are necessary to promote. Progressive cooling is a key preservation technology for cell banks. However, the cell solution needs to be supercooled in a slow freezing process. The high degree of supercooling possibly induces uncontrollable intracellular ice formation. This work designs an ultrasonic ice-seeding system for L-02 hepatocyte cell preservation, reducing supercooling and improving cell survival rate. The effect of ultrasonic intensities on the hepatocyte’s survival rate was investigated and optimized. The results prove the calorimetric method can efficiently measure the ultrasonic intensity dissipated in the hepatocyte cell preservation solution. When the ultrasonic intensity is 0.0329 W/cm2 ∼ 0.4316 W/cm2, the hepatocyte survival rate is over 90%. There is no significant difference between experiment groups (p < 0.05) when the ultrasonic intensity is larger than 0.4316 W/cm2. The hepatocyte cell survival rate reduced significantly with the increase of ultrasonic intensity. The 7-day hepatic function indicator experiment results indicate that the ultrasonic ice seeding has the weakest impact on hepatocyte cells in the four groups. The secretion of urea, albumin and glucose proved that ultrasonic ice seeding technology does not affect cell secretion and has an enormous advantage in cryopreservation. It can be widely applied to cell freezing fields.