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Methodology for Detecting Trace Amounts of Microchimeric DNA from Peripheral Murine White Blood Cells by Real-Time PCR
Journal article   Open access   Peer reviewed

Methodology for Detecting Trace Amounts of Microchimeric DNA from Peripheral Murine White Blood Cells by Real-Time PCR

Carol M. Artlett, C Gennaro Dito and Paul J. Christner
Biological procedures online, v 5(1)
2003
DOI: https://doi.org/10.1251/bpo51
PMID: 12734552
url
https://link.springer.com/content/pdf/10.1251%2Fbpo51.pdfView
Published, Version of Record (VoR) Open
url
https://doi.org/10.1251/bpo51View
Published, Version of Record (VoR) Open

Abstract

Real-time PCR methodology can successfully quantitate microchimeric cell populations at a concentration of 100 microchimeric cells/100,000 host cells; however, it has not been successful in quantitating DNA from trace numbers of microchimeric white blood cells which we reported are present in murine peripheral blood at a concentration as low as 2/100,000 host cells. We report methodology using primers for a portion of the H2-k(b) murine histocompatibility sequence, specific for the C57BL/6J mouse. When these primers were used in the presence of 11,000 microM primer, a 20-fold increase in the median manufacturer's recommended concentration, the assay could be optimized to detect 34 pg of C57BL/6J DNA in a background of 2.5 microg of carrier BALB/cJ DNA (1/100,000). These conditions resulted in a detection limit half as sensitive as that found when no carrier DNA was present.

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