Views: 332 Author: Site Editor Publish Time: 2024-10-26 Origin: Site
Yingtai: Application of Vacuum Freeze-Drying Technology in Cell Biology
Biological research and clinical practices often require the use of red blood cells. However, the collection process for blood red cells can be cumbersome, and opportunities to collect precious blood samples may be limited. Therefore, effective preservation solutions are needed to maintain the viability of the collected red blood cells. Currently, most blood is stored at 4°C, with a shelf life of only about 20 days, which is insufficient for long-term preservation. Additionally, the preservation solutions for red blood cells typically involve short-term storage at room temperature or long-term storage at very low temperatures, leading to significant difficulties in preservation and transport. Furthermore, as storage time increases, red blood cells are prone to oxidative damage, resulting in changes to their membrane structure and function, which subsequently affect their stability and functionality.
1.Application Background
In recent years, freeze-drying technology has been predominantly used for the preservation of proteins and red blood cells. This technology, through the control of low temperatures and vacuum conditions, prevents damage to heat-sensitive and easily oxidizable components in proteins and red blood cells. It also inhibits microbial growth and biochemical reactions mediated by water, thus allowing proteins and red blood cells to retain their original biological activity. Freeze-dried red blood cells can be stored at room temperature for extended periods, maintaining stability and facilitating easy transport and infusion, presenting significant application value in new medical fields. However, during the freeze-drying and rehydration processes, red blood cells are easily affected by external factors, leading to lower survival rates upon rehydration. Furthermore, the currently developed cryoprotectants have complex compositions and difficult-to-control freeze-drying process parameters, resulting in low survival rates, morphological changes, and reduced enzyme activity in cryopreserved red blood cells. Therefore, finding a high-safety, high-survival-rate, and high-activity freeze-drying and rehydration process for red blood cells is crucial for promoting their clinical use.
2. Factors Affecting Drying Rate
Recent studies have shown that the presence of organic solvents can influence the freezing characteristics of solutions, potentially affecting the drying rate and appearance of the freeze-dried products. Tert-butanol, as an organic solvent, has garnered attention from many researchers, but its impact on the freeze-drying process of red blood cells and its effect on recovery rates have not been reported. Experts indicate that the combined use of permeable and non-permeable protective agents can achieve better dehydration before freezing. By using 40% glycerol as a permeable protective agent for pre-treatment of red blood cells, and then adding a non-permeable cryoprotectant for freeze-drying, the study investigates the influence of permeable protective agent pre-treatment on the freeze-drying efficacy of red blood cells. Non-destructive micro-CT scanning technology is utilized to observe red blood cell samples during the sublimation process, aiming to explore the effects of tert-butanol on the drying process. By scanning the freeze-dried red blood cell samples with CT, the porosity can be calculated to determine the impact of glycerol on cell volume and the effect of tert-butanol concentration on ice crystal size. Using a hemocytometer for cell counting before and after freeze-drying allows assessment of the combined effects of glycerol and tert-butanol. During the freeze-drying of red blood cells, micro-CT scanning technology enables timed observations of the sublimation drying process and structural analysis of the freeze-dried products, as well as calculations of porosity values for each sample. For the rehydrated samples, a three-step method is employed for rehydration, with detection using a hemocytometer. Experimental results indicate that the organic solvent tert-butanol can accelerate the freeze-drying process and shorten drying time, with a significant difference observed at a concentration of 10% compared to 5%. During the sublimation process, porosity showed a decreasing trend. The highest porosity was found in the formulation containing 10% tert-butanol with glycerol pre-treatment. Glycerol pre-treatment significantly improved cell recovery rates, while the addition of tert-butanol resulted in a certain degree of decline in recovery rates.