Enhanced ascertainment of microchimerism with real-time quantitative polymerase chain reaction amplification of insertion-deletion polymorphisms

Tzong Hae Lee, Daniel M. Chafets, William Reed, Li Wen, Yunting Yang, Jennifer Chen, Garth H. Utter, John T. Owings, Michael P. Busch

Research output: Contribution to journalArticle

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Abstract

BACKGROUND: The characterization of microchimerism (MC) by gene amplification has been limited by few allogeneic markers, ascertainment bias, and assay analytic performance. To address this, a panel of 12 MC assays based on insertion-deletion (InDel) polymorphisms had been optimized. STUDY DESIGN AND METHODS: The InDel assays were validated with comprehensive in vitro spiking studies at the stochastic limit of detection. Their ability was also determined to ascertain MC of unknown source genotype with both theoretical and actual donor-recipient pairs, and the assays were applied to a clinical population of 73 trauma patients who received transfusions where MC was previously characterized by HLA-based assays alone. RESULTS: In the stochastic spiking experiments, all assays were sensitive to a single copy of target DNA, and no false-positive amplification occurred among 1128 samples studied. Among 219 theoretical donor-recipient pairs, informative alleles existed for 99.5 percent with both InDel and HLA compared to 91.3 percent with HLA alone. In the clinical population, 33 cases of MC were detected (9 more cases than by HLA-DR alone) in the nonleukoreduced (non-LR) group and 8 cases (1 more case than by HLA-DR) in the LR group for the short-term follow-up. Among 27 long-term follow-up samples, 8 cases were detected overall (3 more cases than by HLA-DR alone). CONCLUSION: It is concluded that an InDel-based assay panel has excellent technical performance characteristics while also allowing for ascertainment of some MC cases not detectable with HLA alone. The tandem use of both the InDel and the HLA provides a powerful tool for the enhanced ascertainment of MC.

Original languageEnglish (US)
Pages (from-to)1870-1878
Number of pages9
JournalTransfusion
Volume46
Issue number11
DOIs
StatePublished - Nov 2006

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Chimerism
Real-Time Polymerase Chain Reaction
HLA-DR Antigens
Tissue Donors
Gene Amplification
Population
Limit of Detection
Alleles
Genotype
DNA
Wounds and Injuries

ASJC Scopus subject areas

  • Hematology
  • Immunology

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Enhanced ascertainment of microchimerism with real-time quantitative polymerase chain reaction amplification of insertion-deletion polymorphisms. / Lee, Tzong Hae; Chafets, Daniel M.; Reed, William; Wen, Li; Yang, Yunting; Chen, Jennifer; Utter, Garth H.; Owings, John T.; Busch, Michael P.

In: Transfusion, Vol. 46, No. 11, 11.2006, p. 1870-1878.

Research output: Contribution to journalArticle

Lee, Tzong Hae ; Chafets, Daniel M. ; Reed, William ; Wen, Li ; Yang, Yunting ; Chen, Jennifer ; Utter, Garth H. ; Owings, John T. ; Busch, Michael P. / Enhanced ascertainment of microchimerism with real-time quantitative polymerase chain reaction amplification of insertion-deletion polymorphisms. In: Transfusion. 2006 ; Vol. 46, No. 11. pp. 1870-1878.
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AU - Wen, Li

AU - Yang, Yunting

AU - Chen, Jennifer

AU - Utter, Garth H.

AU - Owings, John T.

AU - Busch, Michael P.

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