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Abstract Background The hemolytic nature of hemolytic disease of the newborn (HDN) is described as the abnormal destruction and decomposition of red blood cells, causing heterogeneous manifestations such as abnormal red blood cell volume and morphology. Mean corpuscular volume (MCV) and red blood cell volume distribution width (RDW) are commonly used parameters related to red blood cell volume. Total serum bilirubin (TSB) is routinely monitored among newborns. This study aims to explore the value of MCV and RDW, combined with TSB, to improve the efficiency of HDN diagnosis. Methods Three hundred eighty-eight children with HDN and 371 children with non-HDN pathological jaundice who were diagnosed and treated in the neonatal department of our hospital from January 2019 to December 2020 were included in the study. Clinical data collected include examination results of laboratory indicators, such as MCV, coefficient of variation of red blood cell volume distribution width (RDW-CV), standard deviation of red blood cell volume distribution width (RDW-SD), and TSB. The differences in the indicators between the two groups of children were retrospectively analyzed. Results 1) The detection rate of HDN in children in the early group was higher than that in the late group (P 163.3 μmol/L, MCV > 96.35fL, and RDW-CV > 16.05%, the diagnosis rate of HDN increased (P 96.35fL or RDW-CV > 16.05%, children with jaundice in three days of birth (especially children with TSB > 163.3 μmol/L) should be screened for HDN. A combination of TSB, MCV, and RDW-CV can improve the early detection rate of HDN, contribute to reduce the readmission rate and risk of hyperbilirubinemia.