A rapid combined immunocytochemical and fluorescence in situ hybridisation method for the identification of human fetal nucleated red blood cells

Abstract

Fetal nucleated red blood cells are found in the maternal circulation during pregnancy. If a simple routine method of detection of these cells was developed, it could be used as the basis of non-invasive prenatal diagnosis of fetal genetic disorders. Fetal male and adult female blood were mixed to mimic maternal blood in pregnancy and used to establish a simple technique to unequivocally detect fetal nucleated red blood cells. These were identified by combined immunocytochemistry using a human fetal haemoglobin antibody and a rapid and simple-to-use fluorescence in situ hybridisation method using X and Y chromosome probes. Initial studies using the alkaline phosphatase anti-alkaline phosphatase technique as the first procedure showed that the stain was unstable and unsuitable for in situ hybridisation. An immunoperoxidase technique was found to produce a stable stain resistant to harsh fixation steps required in subsequent in situ hybridisation. This enabled the simultaneous visualisation of immunopositivity and in situ hybridisation signals on the same cell with neither procedure affecting the other's signal quality. We are currently using this procedure to detect a range of endoplasmic reticulum proteins in fetal nucleated red blood cells from maternal blood in an attempt to diagnose disorders of liver protein expression in early pregnancy.