PubMed · 6083120
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J Wright. 1967. Variations on a theme by Coombs.. https://pubmed.ncbi.nlm.nih.gov/6083120/
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In haemolytic disease of newborn (erythroblastosis fetalis) the in vivo behaviour of erythrocytic IgG antibodies is of particular significance. The monocyte monolayer assay (MMA), which determines by microscopy the number of erythrocytes phagocytosed by monocytes, is an important functional test for the qualitative assessment of erythrocytic IgG antibodies. We set up a photometric MMA and compared it with the microscopic MMA. In both tests we employed commercially available rhesus antibody sera and examined 8 sera of pregnant women with mild and severe courses of haemolytic disease of newborn by the photometric MMA. Good agreement was found between the microscopic and photometric MMA (r = 0.93). Over and above this the photometric MMA correlated with the course of haemolytic disease of newborn in 7 out of 8 cases (with one false-positive finding). The photometric technique permits rapid, sensitive and reproducible determination of erythrophagocytosis in microtitre plates. This method is based on the photometric detection of haemoglobin of phagocytosed erythrocytes via a peroxidase reaction. Standardised photometric MMA could in future be more widely applied especially in haemolytic disease of newborn for the functional characterisation of erythrocytic antibodies, the incidence of intra-assay and inter-assay errors being low.
OBJECTIVE: Current management protocols for pregnancies complicated by red blood cell alloimmunization use the maternal antibody titer to predict the need for invasive testing for detection of fetal anemia. We investigated the use of three maternal serum tests to assess their usefulness in predicting fetal disease: indirect Coombs' titer, Marsh score, and monocyte monolayer assay. STUDY DESIGN: Forty-seven serum samples from pregnant women with red blood cell antibodies associated with fetal anemia were analyzed at cordocentesis. Fetal blood was analyzed for hematocrit (corrected for gestational age) and antigen status. Fetal anemia was defined as a hematocrit value of < 2 SD from the mean value for gestational age. Fetuses were classified into three groups: Antigen positive with anemia (n = 19), antigen positive without anemia (n = 17), antigen negative (n = 11). Statistical methods included Kruskal-Wallis test, Newman-Keuls test, Spearman's rank correlation, and receiver-operator characteristic curves; p < 0.05 was considered significant. RESULTS: The median monocyte monolayer assay (phagocytosis, adherence, and association) did not differ among the three groups. Both maternal titers and Marsh scores were significantly higher in fetuses with anemia compared with the other two groups of fetuses (256 vs 64 vs 64, p < 0.001, and 86 vs 69 vs 64, p = 0.02, respectively). Both titer and Marsh score exhibited significant correlations with corrected fetal hematocrit (r = -0.70, p < 0.001; r = -0.63, p < 0.001, respectively). Comparison of the overall receiver-operator characteristic curves for titer and Marsh score revealed no statistical difference; however, a Marsh score of 57 was noted to have a superior specificity than a titer of 16 (p = 0.02). CONCLUSION: The maternal Marsh score can be performed in conjunction with standard indirect Coombs' titers to enhance the predictability of fetal anemia.
Typically the serological diagnosis of alloimmune haemolytic disease of the newborn (HDN) includes a positive direct antiglobulin test on the infant's red cells, and the presence of an IgG red cell alloantibody in both maternal and cord sera. HDN with a negative direct antiglobulin test has been reported with anti-A and anti-B, but not with other red-cell alloantibodies. In this report we describe four examples of HDN in infants whose red cells had a negative direct antiglobulin test. The first case was diagnosed retrospectively when the infant was admitted to hospital aged 3 weeks with severe anaemia and cardiac failure, and subsequently died. Maternal and infant sera were both shown to contain anti-C: however, the direct antiglobulin test on the infant's red cells was negative. Approximately 1 year later the mother of this infant gave birth to triplets: soon after birth one of the triplets required an exchange transfusion, one had hyperbilirubinaemia, and the third was unaffected. Anti-C and anti-e were detectable in the maternal serum at this time. The most probable Rh genotypes of the two affected infants were R1R2 (CDe/cDE), while the Rh genotype of the unaffected infant was R2R2 (cDE/cDE). Anti-c was implicated as causing HDN in a fourth infant (from a different family) who was a hydropic stillborn. The direct antiglobulin test on fetal blood was negative and other causes of non-immune hydrops were excluded. These four infants provide evidence that the direct antiglobulin test may be negative in some severely affected and even fatal cases of HDN.