Detection of clonal T-cell receptor-gamma gene rearrangement by PCR/temporal temperature gradient gel electrophoresis.
Limited combinatorial and junctional diversity in TCR-gamma gene rearrangement can result in amplification products that are difficult to interpret when analyzed by conventional gel electrophoresis methods that separate DNA based on size (polymerase chain reaction [PCR]/polyacrylamide gel electrophoresis [PAGE]). We describe a simple approach to the detection of clonal TCR-gamma gene rearrangement using temporal temperature gradient gel electrophoresis (TTGE) that uses a gradual and uniform increase in the temperature of a constant denaturing gel to resolve different DNA molecules based on base pair composition. We tested 42 clinical specimens (30 blood specimens and 12 formalin-fixed paraffin-embedded tissues) for T-cell clonality by PCR/PAGE and PCR/TTGE. Concordant results were obtained in only 22 specimens (52%). Of the 20 discordant cases, 18 samples were positive by TTGE and negative by PAGE. For all of the discordant cases, the TTGE yielded results that correlated better with the clinical data than did the PAGE method. We conclude that PCR/TTGE is more accurate and easier to perform than current methods for detecting clonal populations of T cells.
Zhu
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D
Nichols Institute, Quest Diagnostics, 33608 Ortega Highway, San Juan Capistrano, CA, USA.
Kadin
M E
ME
Samoszuk
M
M
D001482:Base Composition; D020029:Base Pairing; D004247:DNA / Q000032:analysis / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D015334:Gene Rearrangement, gamma-Chain T-Cell Antigen Receptor / Q000235:genetics*; D006801:Humans; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013696:Temperature
D004247:DNA