激光显微切割的骨髓巨核细胞中的基因表达谱分析。

Gene expression profiling in laser-microdissected bone marrow megakaryocytes.

作者信息

Hussein Kais

机构信息

Institute of Pathology, Hannover Medical School, Hannover, Germany.

出版信息

Methods Mol Biol. 2011;755:429-39. doi: 10.1007/978-1-61779-163-5_36.

DOI:10.1007/978-1-61779-163-5_36

PMID:21761325

Abstract

Gene expression analysis of the megakaryocytic lineage requires isolation of megakaryocytes from their bone marrow microenvironment. Laser microdissection of megakaryocytes from diagnostic bone marrow biopsies allows analysis of standardised formalin-fixed samples that reflect the in situ grown status quo of a physiological or pathological condition. Taking into account that in neoplastic proliferation, e.g. myeloproliferative neoplasms, non-neoplastic haematopoietic clones proliferate in parallel, this morphology-based isolation enables selective analysis of the aberrant megakaryocytic population. Two different laser microdissection devices are presented, and the details of RNA extraction and subsequent real-time qPCR gene expression analysis of mRNA and microRNA are provided.

摘要

巨核细胞系的基因表达分析需要将巨核细胞从其骨髓微环境中分离出来。对诊断性骨髓活检中的巨核细胞进行激光显微切割，能够分析标准化的福尔马林固定样本，这些样本反映了生理或病理状况下原位生长的现状。考虑到在肿瘤增殖（如骨髓增殖性肿瘤）中，非肿瘤性造血克隆会并行增殖，这种基于形态学的分离方法能够对异常巨核细胞群体进行选择性分析。本文介绍了两种不同的激光显微切割设备，并提供了RNA提取以及随后对mRNA和微小RNA进行实时定量PCR基因表达分析的详细方法。