Laser capture microdissection in pathology.

Abstract

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.