Alternatives to the Kleihauer Betke test: diagnostic performance of three methods in the detection of feto-maternal hemorrhage


      To compare the efficacy of optical fluorescence (OF), flow cytometry (FC), and Kleihauer Betke (KB) in the quantification of fetal-maternal hemorrhage.

      Study design

      Fetal blood was obtained from discarded cord blood of single fetuses delivered before 30 weeks' gestation. Adult blood was obtained from a single male fetal hemoglobin–negative volunteer. OF (using Abbott Celldyn 4000) FC, and KB were compared for their accuracy in discriminating between known concentrations of fetal blood. KB slides were coded so that microscopic counting would be blinded. Precision was determined by performance of multiple replications on each sample. Statistical analyses included Student t, Bland-Altman test for bias, correlation study and precision as determined by linear regression, and Pearson product-moment correlation.


      63 samples were analyzed by FC, OF, and KB. Significant differences between sample means were only present at concentrations <.1% with OF (P<.05 ANOVA) when compared to FC and KB. All testing methods overestimated the 0.02% concentration of fetal cells. Test results from each of the three modalities had a linear relationship. However, no significant difference in accuracy was detected by Bland-Altman testing utilizing sample deviation on a logarithmic scale, or by Pearson's correlation coefficient. A high degree of accuracy and agreement was present on linear regression for all testing methods for the range of 0.1% to 1.0%. There was greater test variation with the KB test at higher concentrations compared to FC or OF.


      Fetal blood cells can be detected on a automated hematological cell analyzer that utilizes fluorescence laser technology. In the future, this technology may allow for the development of a fully automatic method of analyzing maternal fetal hemorrhage.