Detection of weak D and D-VI red cells in D-negative mixtures by flow cytometry: implications for feto-maternal haemorrhage quantification and D typing policies for newborns

Quantitation of fete-maternal haemorrhage (FMH) by flow cytometry (FC) has been shown to be more accurate than the Kleihauer-Bekte test, Fetal cells w ill be predominately of R(1)r or R(2)r phenotype, with antigen site numbers per cell (SPC of between 9900 and 16 000. If the fetus is of weak D or par tial D-VI phenotype, fewer SPC will be present. Red cells from 20 adult weak D samples were mixed with rr red cells to give 1% mixes. Mixtures were stained and analysed by FC, using two different mo noclonal reagents. The SPC of each sample was measured using SOL-ELISA with Scatchard plot analysis. 18 samples could not be distinguished and had <10 00 SPC. Two samples that could be distinguished had 1350 and 3000 SPC. Red cells from seven samples of D-VI were also analysed. None of these samp les could be distinguished: SPC were all <1000. Although one of the reagent s used reacts with D-VI cells, quantitation of a D-VI FMH would not be poss ible due to low SPC, The ability of fetal red cells with low Rh D SPC to cause immunization is q uestionable: failure to measure FMH in these cases is unlikely to cause cli nical problems, as long as suitably sensitive serological reagents and tech niques are used to type all weak D and D variant babies as Rh D positive, a nd thus ensure that the mother is given the appropriate dose of anti-D.