Laser capture microdissection in pathology
Laser capture microdissection in pathology
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DOI：
10.1136/jcp.53.9.666
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发表时间：
2000-09-01
影响因子：
3.4
通讯作者：
Raffeld, M
中科院分区：
文献类型：
--
作者：
Fend, F;Raffeld, M
在DNA、RNA和蛋白质水平上对病理改变的细胞和组织进行分子检测，彻底改变了病理学的研究和诊断。然而，原发组织的固有异质性与各种反应性细胞群的混合物可能会影响分子研究的结果和解释。最近，显微解剖的组织切片和细胞学制备已越来越多地用于分离同质的，形态学鉴定的细胞群，从而克服了组织复杂性的障碍。结合敏感的分析技术，如聚合酶链反应，显微切割允许精确的细胞群，如原位癌或霍奇金病的恶性细胞，这是传统的分子研究无法获得的体内检查。然而，大多数显微切割技术非常耗时，并且需要高度的手动灵活性，这限制了它们的实际应用。激光捕获显微切割（LCM）是美国国家癌症研究所开发的一种新技术，在显微切割的速度、易用性和多功能性方面取得了重要进展。LCM是基于视觉上选择的细胞粘附到热塑性膜上，热塑性膜覆盖在脱水组织切片上，并通过触发低能量红外激光脉冲而局部熔化。熔化的膜与选定的组织区域形成复合物，其可以通过简单地提起膜来去除。LCM可应用于广泛的细胞和组织制备，包括石蜡包埋材料。免疫组织化学染色的使用允许根据表型和功能特征选择细胞。根据起始材料的不同，可以从捕获的组织碎片中成功提取DNA、高质量的mRNA和蛋白质，直至单细胞水平。LCM与表达文库构建、cDNA阵列杂交和差异显示等技术相结合，将建立特异性病理病变，特别是恶性肿瘤的"遗传指纹"。除了识别新的诊断和预后标志物外，这种方法还有助于建立针对肿瘤分子特征的个性化治疗。本文综述了LCM技术，总结了当前的应用和新的方法，并试图给出一个未来的发展前景。此外，LCM与其他最近开发的激光显微切割技术进行了比较。
The molecular examination of pathologically altered cells and tissues at the DNA, RNA, and protein level has revolutionised research and diagnostics in pathology. However, the inherent heterogeneity of primary tissues with an admixture of various reactive cell populations can affect the outcome and interpretation of molecular studies. Recently, microdissection of tissue sections and cytological preparations has been used increasingly for the isolation of homogeneous, morphologically identified cell populations, thus overcoming the obstacle of tissue complexity. In conjunction with sensitive analytical techniques, such as the polymerase chain reaction, microdissection allows precise in vivo examination of cell populations, such as carcinoma in situ or the malignant cells of Hodgkin's disease, which are otherwise inaccessible for conventional molecular studies. However, most microdissection techniques are very time consuming and require a high degree of manual dexterity, which limits their practical use. Laser capture microdissection (LCM), a novel technique developed at the National Cancer Institute, is an important advance in terms of speed, ease of use, and versatility of microdissection. LCM is based on the adherence of visually selected cells to a thermoplastic membrane, which overlies the dehydrated tissue section and is focally melted by triggering of a low energy infrared laser pulse. The melted membrane forms a composite with the selected tissue area, which can be removed by simple lifting of the membrane. LCM can be applied to a wide range of cell and tissue preparations including paraffin wax embedded material. The use of immunohistochemical stains allows the selection of cells according to phenotypic and functional characteristics. Depending on the starting material, DNA, good quality mRNA, and proteins can be extracted successfully from captured tissue fragments, down to the single cell level. In combination with techniques Like expression library construction, cDNA array hybridisation and differential display, LCM will allow the establishment of "genetic fingerprints" of specific pathological lesions, especially malignant neoplasms. In addition to the identification of new diagnostic and prognostic markers, this approach could help in establishing individualised treatments tailored to the molecular profile of a tumour. This review provides an overview of the technique of LCM, summarises current applications and new methodical approaches, and tries to give a perspective on future developments. In addition, LCM is compared with other recently developed laser microdissection techniques.