Published online before print July 9, 2007

This Article Full Text (Rapid PDF) All Versions of this Article: jmoldx.2007.070006v1 9/4/490    most recent Purchase Article View Shopping Cart Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Yakirevich, E. Articles by Resnick, M. B. Search for Related Content PubMed PubMed Citation Articles by Yakirevich, E. Articles by Resnick, M. B.

Copyright © 2007 American Society for Investigative Pathology
Journal of Molecular Diagnostics, doi:10.2353/jmoldx.2007.070006

Accepted for publication May 16, 2007.

Article

Analysis of T-Cell Clonality Using Laser Capture Microdissection and High-Resolution Microcapillary Electrophoresis

From the Department of Pathology, Rhode Island Hospital and Brown Medical School, Providence, Rhode Island

^@ To whom correspondence should be addressed. E-mail: eyakirevich{at}lifespan.org.

	Abstract

Identification of clonal lymphocytic populations by polymerase chain reaction may be difficult in cases with scant cellular infiltrates or those with a heterogeneous population of cells. Here, we assessed the diagnostic utility of laser capture microdissection (LCM) and high-resolution microcapillary electrophoresis in the analysis of clonality of small biopsy specimens. Clonality was determined in 24 cases: five reactive tonsils, five reactive lymph nodes, six inflammatory skin lesions, and eight T-cell lymphomas. CD3-positive T lymphocytes were captured by LCM from paraffinized immunohistochemically stained sections. Genomic DNA was analyzed for T-cell receptor- gene rearrangement by polymerase chain reaction followed by high-resolution microcapillary electrophoresis with the DNA 500 LabChip and the Agilent Bioanalyzer. In the reactive specimens, T-cell receptor- polymerase chain reaction revealed monoclonal bands when 10 to 1000 cells were captured. This pattern changed to polyclonal when higher numbers of cells were microdissected (2000 to 10,000 cells). In contrast, lymphoma cells were consistently monoclonal whether low or high numbers were microdissected. Microcapillary electrophoresis coupled with LCM facilitated clonality analysis in equivocal cases. In two of eight lymphoma cases, LCM revealed diagnostic monoclonal bands, whereas routine T-cell receptor- assessment of whole tissue sections with 10% polyacrylamide gel electrophoresis demonstrated only minor clonal bands. We conclude that clonality determined by LCM is cell number-dependent. Biopsy specimens containing low numbers of reactive polyclonal T cells may produce pseudomonoclonal bands and therefore should be interpreted with great caution.