Quantification of male (foetal) cells in a background of female (maternal) cells.
As part of a research program aimed at isolating foetal cells from maternal blood samples we have developed methodologies to enumerate low numbers of foetal cells in a background of maternal blood cells. Using a model system constructed using male cells flow sorted into a high background of female cells, we have optimised a DNA extraction method and combined this with real-time quantitative PCR to determine the accuracy of quantifying low numbers of male cells in a high background of female cells. Male cells (between 1 - 1000) were sorted into tubes containing approximately 100,000 female cells. DNA was extracted and quantitation was carried out using b-globin primers (for total cell numbers) and tspy primers (a multicopy Y chromosome-specific target), utilising a standard curve created with male genomic DNA. Approximately 50% of input cells could be identified when between 50 and 1000 male cells were added. Accurate quantitation was not possible for 1 - 10 male cells, although male DNA was detected in 100% of samples containing male cells.