Laser capture microdissection in the tissue biorepository

Abstract

An important need of many cancer research projects is the availability of high-quality, appropriately selected tissue. Tissue biorepositories are organized to collect, process, store, and distribute samples of tumor and normal tissue for further use in fundamental and translational cancer research. This, in turn, provides investigators with an invaluable resource of appropriately examined and characterized tissue specimens and linked patient information. Human tissues, in particular, tumor tissues, are complex structures composed of heterogeneous mixtures of morphologically and functionally distinct cell types. It is essential to analyze specific cell types to identify and define accurately the biologically important processes in pathologic lesions. Laser capture microdissection (LCM) is state-of-the-art technology that provides the scientific community with a rapid and reliable method to isolate a homogeneous population of cells from heterogeneous tissue specimens, thus providing investigators with the ability to analyze DNA, RNA, and protein accurately from pure populations of cells. This is particularly well-suited for tumor cell isolation, which can be captured from complex tissue samples. The combination of LCM and a tissue biorepository offers a comprehensive means by which researchers can use valuable human biospecimens and cutting-edge technology to facilitate basic, translational, and clinical research. This review provides an overview of LCM technology with an emphasis on the applications of LCM in the setting of a tissue biorepository, based on the author's extensive experience in LCM procedures acquired at Fox Chase Cancer Center and Hollings Cancer Center.

Keywords: cancer biology; cells of interest; pathology.

FIGURE 1

The workflow of LCM. Tissue sections were cut using a cryostat or microtome and stained with hematoxylin and eosin (H&E) for cell identification. Pure epithelial cell populations from frozen ovarian tissue were captured using the Arcturus XT LCM system and visualized on the LCM cap. Total RNA was then extracted, and the quantity and quality of RNA were monitored on an Agilent bioanalyzer. Two peaks, 18S and 28S ribosomal RNA, were obtained in the RNA profile generated by the bioanalyzer, indicating successful RNA sample isolation. The RNA was amplified for downstream analysis. QC, quality control; qPCR, quantitative PCR.

FIGURE 2

RNA profile generated by Agilent Bioanalyzer. High-quality RNA was extracted from LCM-isolated cells. RNA size and quality were analyzed with the Agilent 2100 bioanalyzer. A pseudo gel image was created, and bands were sized and quantified. (A and B) The y-axes indicate fluorescence units (FU), and the x-axes indicate the length of the RNA in nucleotides. The peaks of 18S and 28S rRNA are clearly visible. Shown are the profiles of total RNA extracted from scrapes of the entire prostate section (A) and from LCM-isolated cells (B). (C) Gel electrophoresis analysis of the same RNA in A and B.

FIGURE 3

Capture of pure cell populations by LCM. (A) LCM of HL and RS cells. A HL tissue section was stained with H&E (left), and the giant RS cells were captured successfully by LCM and visualized on the LCM cap (middle). (B) LCM of pure cell populations from a prostate cancer sample. A prostate adenocarcinoma tissue section was stained with H&E (left), and the benign, prostatic intraepithelial neoplasia (PIN), malignant, and stromal cells were isolated by LCM, respectively, and visualized on the LCM cap (middle). All human tissues were obtained from patients who provided informed consent and acquired through the Hollings Cancer Center Tissue Biorepository in accordance with an Institutional Review Board-approved protocol.