Investigation of the correlation between immature reticulocyte fraction and reticulocyte hemoglobin content with common tests for iron deficiency anemia

Reticulocyte hemoglobin content (Ret-He, the hemoglobin within reticulocytes or immature red blood cells) and immature reticulocyte fraction (IRF, the immature fraction of the absolute-reticulocyte-count) are tests that provide insight into erythropoiesis and iron status earlier than conventional iron studies offering the added benefit of not being acute-phase-reactants. Studies have shown that Ret-He is a diagnostic marker for iron-deficiency-anemia (IDA), but fewer studies have investigated IRF. Our laboratory is currently planning to report these parameters when reticulocyte is ordered. Since these are new parameters, we wanted to investigate their overall correlation with complete blood count (CBC) and other iron studies to gain a better appreciation of their utility in our patient population. The aim of this study was to compare the overall correlation of Ret-He and IRF with seven tests used in the evaluation of IDA. To our knowledge these parameters have not all been directly correlated within a single study. CBC and reticulocytes were quantified using XN 9000 hematology analyzers (Sysmex Corporation), ferritin (DXI 800, Beckman Coulter Inc.), and % iron-saturation (measured using total iron-binding-capacity (TIBC)=transferrin*1.18 on Cobas 6000, Roche Diagnostics). Two de-identified cohorts of patients undergoing physician-ordered reticulocyte testing were used for this analysis. Dataset 1 (DS1): (N=2026 from Mayo Clinic Florida) had Ret-He and IRF compared to absolute-reticulocyte-count (Ret), ferritin and % iron saturation. Dataset 2 (DS2): (N=3990 from Mayo Clinic Rochester) had Ret-He and IRF compared to the red-cell-indices of the CBC including hemoglobin (Hgb), mean-corpuscular-volume (MCV), mean-corpuscular-hemoglobin (MCH), and mean-corpuscular-hemoglobin-concentration (MCHC). Correlation coefficients were calculated using Spearman rank-order (ρ) wherein values below +/-0.39 are weak, between +/-0.40-0.59 are considered moderate, and values above +/-0.60 are considered strong. For DS1, Ret-He demonstrated the following correlations: Ret (ρ=0.01), ferritin (ρ=0.33), % iron saturation (ρ=0.63). IRF demonstrated: Ret (ρ=0.46), ferritin (ρ=-0.05), % iron saturation (ρ=-0.22). For DS2, Ret-He demonstrated the following correlations: Hgb (ρ=0.17), MCV (ρ=0.64), MCH (ρ=0.74), MCHC (ρ=0.56). IRF demonstrated Hgb (ρ=-0.41), MCV (ρ=0.10), MCH (ρ=0.04), MCHC (ρ=-0.11). Ret-He and IRF demonstrated different correlative profiles suggesting they may have differing uses. Ret-He was strongly positively-correlated with % iron saturation, MCV, MCH and moderately positively-correlated with MCHC. These positive-correlations are consistent with relationships established in the literature. Interestingly, Ret-He was only weakly correlated with ferritin, possibly owing to ferritin being an acute-phase-reactant. IRF had a moderate positive correlation with Ret and moderate inverse correlation with Hgb. Both of these IRF relationships are consistent with other reports, but both relationships have not been shown in the same study before, preventing direct comparison until now. The literature suggests IRF may have more potential in monitoring treatment than in diagnosis. One limitation of these datasets is their lack of clinical correlation such as established iron-deficiency, anemia status, or treatment information.