使用最佳渗透压和双密度梯度系统分离出高产率的胎儿有核红细胞。

A high yield of fetal nucleated red blood cells isolated using optimal osmolality and a double-density gradient system.

作者信息

Kwon K H, Jeon Y J, Hwang H S, Lee K A, Kim Y J, Chung H W, Pang M G

机构信息

School of Public Health and Institute of Health and Environment, Seoul National University, Seoul, Korea.

出版信息

Prenat Diagn. 2007 Dec;27(13):1245-50. doi: 10.1002/pd.1888.

DOI:10.1002/pd.1888

PMID:17994634

Abstract

OBJECTIVES

To increase the yield of fetal nucleated red blood cells (NRBCs) from maternal blood using a discontinuous Percoll gradient and to determine the effects of osmolality on NRBC yield.

METHODS

Fetal NRBCs were isolated from combined umbilical cord blood and adult female blood, or from maternal blood using single or double Percoll gradients with different osmolalities. Magnetic activated cell sorting was used to enrich isolated NRBCs, and morphological differentiation was performed with Kleihauer-Betke stain. We also isolated fetal NRBCs from 25 10 mL samples of maternal blood and determined fetal sex by fluorescence in situ hybridization (FISH), using X-Y probes.

RESULTS

For single-density Percoll columns, the greatest number of NRBCs was isolated using 280 mOsm/kg H(2)O with 1.077 g/mL Percoll and 520 mOsm/kg H(2)O with 1.119 g/mL Percoll. Significantly more fetal NRBCs were isolated with double Percoll density gradients than with double-Histopaque gradients (p = 0.043). FISH analysis on NRBC in 25 cases correctly identified 15 male and 9 female euploid fetuses and one Trisomy 21 fetus.

CONCLUSION

The NRBC enrichment method we present requires less maternal blood and yields more NRBCs compared to previous methods.

摘要

目的

使用不连续的 Percoll 梯度提高从母血中获取胎儿有核红细胞（NRBCs）的产量，并确定渗透压对 NRBC 产量的影响。

方法

从脐带血与成年女性血液的混合样本中，或使用具有不同渗透压的单 Percoll 梯度或双 Percoll 梯度从母血中分离胎儿 NRBCs。使用磁珠细胞分选技术富集分离出的 NRBCs，并采用 Kleihauer-Betke 染色进行形态学鉴别。我们还从 25 份 10 mL 的母血样本中分离胎儿 NRBCs，并使用 X-Y 探针通过荧光原位杂交（FISH）确定胎儿性别。

结果

对于单密度 Percoll 柱，使用渗透压为 280 mOsm/kg H₂O 且 Percoll 密度为 1.077 g/mL，以及渗透压为 520 mOsm/kg H₂O 且 Percoll 密度为 1.119 g/mL 时，分离出的 NRBCs 数量最多。与双 Histopaque 梯度相比，使用双 Percoll 密度梯度分离出的胎儿 NRBCs 明显更多（p = 0.043）。对 25 例样本中 NRBC 的 FISH 分析正确鉴定出 15 例男性和 9 例女性整倍体胎儿以及 1 例 21 三体胎儿。