Rosette Test

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The page below is a sample from the LabCE course Hemolytic Disease of the Fetus and Newborn (retired 4/6/2020). Access the complete course and earn ASCLS P.A.C.E.-approved continuing education credits by subscribing online.

Learn more about Hemolytic Disease of the Fetus and Newborn (retired 4/6/2020) (online CE course)
Rosette Test

The rosette test is a screening test for FMH that detects fetal D+ red cells in maternal Rh negative blood. If the rosette test is positive, follow-up testing is done to quantitate the FMH, eg, a Kleihauer-Betke acid elution test or flow cytometry.

Note: The rosette test may be falsely positive if the mother is weak-D positive and may be falsely negative if the baby is weak-D positive. In such cases, a Kleihauer-Betke test or flow cytometry should be done.

General description (example only):
  • Incubate a maternal 3-5% red cell suspension with IgG anti-D at 37°C. The anti-D will bind to any infant D+ cells that are present.
  • After washing to remove unbound anti-D, add indicator red cells. Indicator cells are ficin-treated R2R2 cells that will bind to the antibody-coated infant RBCs causing agglutination (“rosettes”) that can be detected microscopically.
  • A specified number of agglutinates (eg, 3 or more in 10 fields or 7 or more in 5 fields) is designated a positive and suggests a significant FMH (>30 mL) requiring more RhIG.
The top image on the right illustrates a negative rosette test. The bottom image is representative of a field that would meet the criteria for a positive rosette test, if the same number of agglutinates, or more, are counted in the required number of fields, as discussed above.
These images were provided courtesy of Mount Sinai Blood Transfusion Laboratory, Toronto, Ontario.