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一种基于胰蛋白酶的方法,用于从人蜕膜中分离白细胞,适用于免疫表型和转录组研究。

A trypsin-based method for isolating leukocytes from human choriodecidua suitable for immunophenotyping and transcriptome studies.

机构信息

Departamento de Nutrición y Bioprogramación, Instituto Nacional de Perinatología Isidro Espinosa de los Reyes, Montes Urales 800, Lomas Virreyes, 11000 Mexico City, Mexico.

Departamento de Nutrición y Bioprogramación, Instituto Nacional de Perinatología Isidro Espinosa de los Reyes, Montes Urales 800, Lomas Virreyes, 11000 Mexico City, Mexico; Departamento de Biomedicina Molecular Centro de Investigación y de Estudios Avanzados, Instituto Politécnico Nacional Av Instituto Politécnico Nacional 2508, La Laguna Ticoman, 07360 Mexico City, Mexico.

出版信息

Immunobiology. 2019 Jan;224(1):177-181. doi: 10.1016/j.imbio.2018.09.001. Epub 2018 Sep 22.

Abstract

Leukocytes found at the human maternal-fetal interface participate in the inflammatory process associated with both preterm and term labor; therein, effective methods for their isolation that allow further phenotypic and functional analyses are necessary. Leukocyte isolation is usually carried out through scraping or enzyme digestion of the choriodecidua, however both methods usually limit the use of downstream immunophenotyping or transcriptomic techniques. Here we describe an isolation method based on gentle trypsin digestion that yields a leukocyte-enriched cell mixture with high lymphocyte viability, although less viable myeloid cells. We show that the method does not compromise cell surface markers since isolated leukocytes are suitable for flow cytometry; and that high quality RNA can be obtained from these cells for qRT-PCR and microarray analyses.

摘要

在人类母体-胎儿界面发现的白细胞参与与早产和足月分娩相关的炎症过程;因此,需要有效的方法将其分离出来,以进行进一步的表型和功能分析。白细胞分离通常通过刮取或酶消化绒毛膜蜕膜来进行,但这两种方法通常限制了下游免疫表型或转录组学技术的应用。在这里,我们描述了一种基于温和胰蛋白酶消化的分离方法,该方法产生富含淋巴细胞的细胞混合物,具有较高的淋巴细胞活力,尽管骨髓细胞的活力较低。我们表明,该方法不会影响细胞表面标志物,因为分离的白细胞适合流式细胞术;并且可以从这些细胞中获得用于 qRT-PCR 和微阵列分析的高质量 RNA。

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