Flow cytometric measurement of HLA-DR expression on circulating monocytes in healthy and sick neonates using monocyte negative selection

The aim of this study was to investigate the effect of prematurity, neonata l sepsis, respiratory distress syndrome (RDS) and perinatal asphyxia on mon ocyte HLA-DR expression of neonates using a flow cytometric method based on monocyte negative selection. The subjects were one hundred and thirty-one neonates (59 healthy, 44 septicaemic, 20 with RDS and eight with perinatal asphyxia) and 20 healthy adults. Monocyte HLA-DR expression was measured us ing one-colour HLA-DR labelling in a gate for monocytes obtained using the combination of CD3-CD19-PE/CD15-FITC MoAbs. In addition, the common dual st aining method using MoAbs against two CD14 epitopes (TUK4, MO2) was evaluat ed. With the one-colour HLA-DR labelling higher purity and recovery values of monocytes were achieved than with the dual labelling method. Healthy neo nates had significantly lower percentages of HLA-DR+ monocytes than adults (69 +/- 13% versus 91.5 +/- 2.5%) and comparable mean fluorescence intensit y (MFI) (119 +/- 25 versus 131 +/- 26). Values did not differ significantly between healthy term and preterm neonates. Preterm neonates with RDS had a significantly lower percentage of HLA-DR+ monocytes than the healthy prete rm neonates. In neonates with asphyxia both parameters were comparable to t hose of the healthy ones. Septicaemic neonates presented significantly lowe r values of both parameters than the healthy, RDS and asphyxiated neonates. Monocyte negative selection provides a reliable estimation of HLA-DR expre ssion on monocytes. Expression of monocyte HLA-DR is lower in healthy neona tes in comparison with adults and is further decreased in neonates with sep sis and RDS, but it is not influenced by prematurity and perinatal asphyxia .