JH probe real‐time quantitative polymerase chain reaction assay for Bcl‐2/IgH rearrangements

Follicular lymphoma (FL) characteristically bears the t(14;18)(q32;q21). However, only ∼75% of the consequent Bcl‐2 breakpoints lie within the major breakpoint region (MBR) or the minor cluster region (mcr). While these can be quantified by cluster region‐specific real‐time quantitative polymerase c...

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Bibliographic Details
Published in:British journal of haematology 2002-08, Vol.118 (2), p.550-558
Main Authors: Jenner, Michael J., Summers, Karin E., Norton, Andrew J., Amess, John A., Arch, Rachael S., Young, Bryan D., Lister, T. Andrew, Fitzgibbon, Jude, Goff, Lindsey K.
Format: Article
Language:English
Subjects:
Bcl‐2
Chromosome Breakage
Chromosomes, Human, Pair 14 - genetics
Chromosomes, Human, Pair 18 - genetics
follicular lymphoma (FL)
Gene Rearrangement - genetics
Genes, bcl-2
Hematology
Humans
IgH
Immunoglobulin Heavy Chains - genetics
Lymphoma, Follicular - genetics
molecular monitoring
Polymerase Chain Reaction
real‐time PCR (RQ‐PCR)
t(14
18)
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Summary:Follicular lymphoma (FL) characteristically bears the t(14;18)(q32;q21). However, only ∼75% of the consequent Bcl‐2 breakpoints lie within the major breakpoint region (MBR) or the minor cluster region (mcr). While these can be quantified by cluster region‐specific real‐time quantitative polymerase chain reaction (RQ‐PCR), a significant proportion of cases are left requiring a customized approach. Therefore, an RQ‐PCR assay for the quantification of Bcl‐2/IgH breakpoints has been developed that uses germline JH TaqMan probes and germline JH primers in combination with customized forward primers. Validation of this approach by comparison with an established MBR RQ‐PCR showed both techniques to be concordant across a wide range of copy numbers with a sensitivity of five copies per 105 cells. In addition, to generate standard curves equating to diverse Bcl‐2/IgH rearrangements, a strategy for using placental DNA as a surrogate standard was devised. The performance of the assay in detecting molecular evidence of disease in sequential biopsies from five patients (three with atypical Bcl‐2/IgH breakpoints identified by long‐range or inverse PCR, one MBR+ and one mcr+) was tested. This alternative approach represents a sensitive and specific means of quantifying common and atypical Bcl‐2/IgH rearrangements and maximizes the number of patients with FL suitable for molecular monitoring.
ISSN:0007-1048
1365-2141
DOI:10.1046/j.1365-2141.2002.03623.x