Comparison of Gene Expression Profiles in Laser-Microdissected, Nonembedded, and OCT-Embedded Tumor Samples by Oligonucleotide Microarray Analysis

In the study and interpretation of expression profiles in tumor samples, the quantity and quality of the RNA and the heterogeneity of the tissue specimen from which it is extracted are both important. Optimization of amplification protocols has allowed researchers to use fewer tumor cells to obtain expression profiles for some clinical samples (1)(2). Assessment of RNA quality control has been improved by the use of highly sensitive gel analysis (3), and flow cytometry and laser microdissection techniques allow isolation of tumor cells from nondiseased connective tissue as well as inflammatory infiltrates (4)(5). Several groups have reported the use of gene expression profiles from laser-microdissected tumors (6)(7)(8). However, the extent of the effect of the laser beam on the quality of the RNA and on the expression profile results remains unclear. Because the amount of RNA obtained after laser microdissection is generally low, opportunities for additional quality control and validation are limited (9)(10). These issues led us to investigate the potential differences in gene expression profiles among samples obtained by different sample manipulation procedures. We compared the gene expression profiles of nonmicrodissected (NE) samples with those from manually microdissected OCT blocks (OCT) and laser capture-microdissected sections (LASER; obtained with a PixCell microdissector) from aliquots of tumors obtained from the same patients. Four bladder transitional cell carcinomas and one breast carcinoma were evaluated. The expression profiles of the 15 tumor samples obtained with oligonucleotide HG-133A arrays containing 22 283 probes (Affymetrix) were analyzed by means of a t -test and hierarchical clustering (11)(12). We estimated the effect of laser microdissection on gene profiling as well as potential sources of differences and criteria that may enable characterization of a sample as suitable for gene expression analysis. A detailed description of …