Real-time sequence-specific primer polymerase chain reaction amplification of HLA class II alleles: A novel approach to analyze microchimerism

Gloria Garavito, Dagmar Klein, Melvin Denis, Alberto Pugliese, Camillo Ricordi, Ricardo L. Pastori

Research output: Contribution to journalArticle

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Abstract

The careful assessment of microchimerism is essential to investigate the effects of donor bone marrow-derived cells in transplantation. We have developed a protocol to assess microchimerism based on the HLA mismatch between the recipient and the donor. Our approach combines real-time polymerase chain reaction (PCR) with sequence-specific primer PCR (SSP-PCR) to selectively amplify and measure the abundance of donor HLA alleles in DNA samples extracted from the recipient after transplant. To optimize and validate the reliability of this method at different levels of microchimerism, we tested serial dilutions of donor DNA into recipient DNA. We demonstrate that donor alleles can be readily detected and reliably measured at concentrations as low as 0.1%. This method is simple and rapid and could find practical application in the assessment of microchimerism in patients receiving organ or cellular transplants in conjunction with donor bone marrow cells infusion.

Original languageEnglish (US)
Pages (from-to)822-825
Number of pages4
JournalTransplantation
Volume73
Issue number5
DOIs
StatePublished - Mar 15 2002

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ASJC Scopus subject areas

  • Transplantation

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