Quantification of all fetal nucleated cells in maternal blood between the 18th and 22nd weeks of pregnancy using molecular cytogenetic techniques

Abstract

Different types of nucleated fetal cells (trophoblasts, erythroblasts, lymphocytes, and granulocytes) have been recovered in maternal peripheral blood. In spite of many attempts to estimate the number of fetal cells in maternal circulation, there is still much controversy concerning this aspect. The numbers obtained vary widely, ranging from 1 nucleated cell per 104 to 1 per 109 nucleated maternal cells. The purpose of our project was to determine the absolute number of all different types of male fetal nucleated cells per unit volume of peripheral maternal blood. Peripheral blood samples were obtained from 12 normal pregnant women known to carry a male fetus between 18 and 22 weeks of pregnancy. Three milliliters (3 ml) of maternal blood has been processed without any enrichment procedures. Fluorescence in situ hybridization (FISH) and primed in situ labeling (PRINS) were performed, and fetal XY cells were identified (among maternal XX cells) and scored by fluorescent microscopy screening. The total number of male fetal nucleated cells per milliliter of maternal blood was consistent in each woman studied and varied from 2 to 6 cells per milliliter within the group of normal pregnancies. The number of fetal cells in maternal blood, at a given period, is reproducible and can therefore be assessed by cytogenetic methods. This confirms the possibility of developing a non-invasive prenatal diagnosis test for aneuploidies. Furthermore, we demonstrate that it is possible to repeatedly identify an extremely small number of fetal cells among millions of maternal cells.

CD34+ cells in maternal placental blood are mainly fetal in origin and express endothelial markers

“…1,2 This phenomenon starts early, as soon as at 6 weeks of gestation in a small proportion of women. 1 The number of fetal cells has been estimated about 1-6 per ml of maternal blood during the second trimester 2 and seems influenced by obstetrical events such as preeclampsia, 3 abortion, 4,5 amniocentesis or fetal abnormalities, such as Down syndrome, 6 each of which result in higher amounts of transferred cells. The phenotype of the fetal cells transferred to the mothers has also been investigated despite their low frequency.…”

The Estimationof Possibility of Isolation and Analysis of Trophoblast Cells in the Model Experiment

“…Было показано, что в 1 мл крови беременной женщины содержится 1-6 клеток плода, и большая часть этих клеток имеет трофобластную природу [4,5]. Крайне низкое содержание плодных клеток в материнском кровотоке является причиной нелегкого поиска оптимального способа выделения клеток плода из образца материнской крови.…”

Efficacy of Primed In Situ Labelling in Determination of HER-2 Gene Amplification and CEN-17 Status in Breast Cancer Tissue

“…An additional improvement was the direct use of fluorochromes in sequential PRINS reactions. A multicolour PRINS protocol has been reported, allowing performance of ultra-rapid detection on several chromosomes, by mixing the different fluorochromes during the chain elongation reaction (Yan et al, 2001). PRINS reactions are fast, and the resulting data can be obtained in less than 4 hrs, whereas FISH results for HER-2 detection are generally obtained at least after 16-20 hrs or even more.…”

“…The use of PRINS has also been reported for analysis of structural aberrations such as translocations and marker chromosomes and localization of single copy genes such as SRY and SOX3 (Kadandale et al, 2000a(Kadandale et al, , 2000b as well as for the detection of fetal cells in peripheral venous blood of pregnant women (Orsetti et al, 1998;Krabchi et al, 2001;Krabchi et al, 2006). Further applications of PRINS have also been reported for tumoral cytogenetics (Tharapel and Kadandale, 2002).…”