Obtaining clone-specific primer and probe for the immunoglobulin heavy- chain gene from paraffin-embedded tissue of B-cell lymphoma: Technical considerations

G. Wu, T. C. Greiner, Wing C. Chan

Research output: Contribution to journalArticle

15 Scopus citations

Abstract

The complementarity determining region (CDR) III of the immunoglobulin heavy-chain (IgH) gene is a tumor-specific marker for B-cell malignancies that has been widely exploited for the monitoring of minimal residual disease in B-precursor acute lymphocytic leukemia. There are a number of technical problems in applying the same technology for B-cell non-Hodgkin's lymphoma (B-NHL). Several procedures have been useful in overcoming these unique problems encountered in obtaining the tumor-specific sequence of the IgH- CDRIII in B-NHL, including the use of denaturing gradient gel electrophoresis or micromanipulation of tissue sections in separating the tumor-specific CDRIII products from those of contaminating normal B-lymphocytes. Minor modifications of a commercial kit greatly improve the purity of the polymerase chain reaction (PCR) products for sequencing. Modifications of the 5'-ends of the V(H) and I(H) primers, coupled with the cycle sequencing technique, make it possible to obtain unambiguous sequences on direct sequencing of short PCR products. Computer informatics and programs that facilitate the design of tumor-specific primers and probes from CDRIII sequences are described.

Original languageEnglish (US)
Pages (from-to)147-153
Number of pages7
JournalDiagnostic Molecular Pathology
Volume6
Issue number3
DOIs
StatePublished - Jun 1997

Keywords

  • B-cell malignancies
  • Complementaritydetermining region
  • DNA sequencing
  • Immunoglobulin heavy-chain gene
  • Minimal residual disease
  • Polymerase chain reaction
  • Tumor-specific primer
  • Tumor-specific probes

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Molecular Biology
  • Cell Biology

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