甘露醇在冷冻解冻去甘油红细胞中应用的研究

Explore the application of mannitol in frozen-thawed deglycerolized red blood cells

目的探讨在红细胞冷冻前、解冻洗涤过程中添加适宜浓度大分子细胞膜保护剂-甘露醇,提高冷冻解冻去甘油红细胞RBC回收率、降低甘油残存量的可行性。方法选取20袋6 d内制备的400 mL去白细胞悬浮红细胞,离心去浆后等量分装(各100 mL)作为实验组和对照组,其中实验组在160 mL甘油化试剂、解冻第1次洗涤80mL 9%NaCl溶液、第2次洗涤200 mL 0.9%NaCl溶液中分别加入终浓度为1.6%、1%、0.2%的甘露醇,通过统计学分析比较实验组与对照组RBC回收率、甘油残存量、红细胞渗透脆性是否有显著性差异,并通过检测冷冻红细胞解冻洗涤过程中游离血红蛋白含量,确定提高冷冻解冻去甘油红细胞RBC回收率的关键点。结果实验组RBC回收率显著高于对照组(t=20.727,P<0.05)、甘油残存量显著低于对照组(t=-4.611,P<0.05)、红细胞渗透脆性与对照组差异无统计学意义(t=0.317,P>0.05),冷冻红细胞解冻第1次洗涤后实验组血浆游离血红蛋白含量低于对照组(t=-22.326,P<0.05),其余洗涤步骤实验组与对照组血浆游离血红蛋白含量差异无统计学意义(P>0.05)。结论冷冻解冻去甘油红细胞在160 mL甘油化试剂、解冻第1次洗涤80 mL 9%NaCl溶液、第2次洗涤200 mL0.9%NaCl溶液中分别加入终浓度为1.6%、1%、0.2%的甘露醇,红细胞回收率显著提高、甘油残存量显著降低、红细胞渗透脆性无显著性差异,冷冻解冻去甘油红细胞RBC回收率提高的关键点为解冻第一次洗涤过程。

Objective To explore the feasibility of improving the recovery ratio of red blood cells and reducing the resi- due level of glycerin in forzen-thawed deglycerolized red blood cells by adding an appropriate amount of macromoleeular mem- brane protectant-mannitol before freezing and during the washing process. Methods 20 packs of 400 mL red blood cells which were leukocyte-removed and prepared within 6 days were selected. The plasma was removed after centrifuge and each pack was divided into 2 bags （ 100mL each） as experimental group and control group. In the experimental group,mannitolwith final concentration of 1.6% ,1% ,0. 2% was added into 160 mL glycerol reagent,80 mL 9% NaCl（first washing solution） and 200 mL 0. 9% NaCI（ second washing solution）. The recovery ratio of red blood cells, residue level of glycerin and osmotic fragility in two groups were analyzed by statistical methods. The key point of improving recovery ratio of red blood cells was sought by detecting the concentrate of free hemoglobin in washing process. Results Among those packs in experimental group, the recovery ratio of red blood cells was significantly higher than that in control group（ t = 20. 727, P 〈 0. 05 ）while the residue level of glycerin and the concentration of free hemoglobin after first washing were significantly lower than those in control group（ t r- - 4. 611, P 〈 0. 05 ; t = - 22. 326, P 〈 0. 05 ）. There were no significant difference on osmotic fragility （ t = 0. 317, P 〉 0. 05 ） and free hemoglobin concantration after other Washing steps （ P 〉 0. 05 ） between experimental group and control group. Conclusion The recovery ratio of red blood ceils could be improved and the residue level of glycerin could be reduced in forzen-thawed deglycerolized red blood cells by respectively adding mannitol with the final concentration of 1.6%, 1% ,0. 2% into 160 mL glycerol reagent,80 mL 9% NaCl（first washing solution） and 200 mL 0. 9% NaCl （second washing solution） ,while the osmotic fragility could stay the same. The key point of improving recovery ratio of forzen-thawed deglycer- olized red blood ceils was the first washing process.