Immature Reticulocyte Research Articles

The importance of immature granulocyte and immature reticulocyte fraction for the severity of acute bronchiolitis

The immature granulocyte (IG) and immature reticulocyte fraction (IRF) are new analytical parameters of the complete blood count, that have been studied as biomarkers of several inflammatory conditions. Here, our aim is to determine the effectiveness of IG and IRF percentages for the severity of acute bronchiolitis (AB). A single-center, prospective study was performed in patients who were hospitalized for acute bronchiolitis and healthy children were included as a control group. The demographic characteristics, white blood cell (WBC) count, platelet (PLT) count, eosinophil%, IG%, and IRF% values were analysed. Receiver operating characteristics (ROC) analysis was used to compare the diagnostic accuracies and predictive performances. We enrolled 168 infants in the acute bronchiolitis group and 80 in the control group. The Clinical Severity Score (CSS) showed that 48, 93, and 27 patients had mild, moderate, and severe bronchiolitis, respectively. The WBC, PLT, and IRF value were significantly higher in patients (p < 0.001). There was no difference between the patients and control group in terms of IG and eosinophil percentage. Only a positive correlation was observed between the clinical severity of the AB and IRF (p=0.003).The ROC curve analysis indicated that the IRF% cut-off point for predicting severity AB was >12.4, with a sensitivity of 53% and specificity of 88% (Areas under the curves (AUC):0.701,p<0.001). The WBC count, PLT count, and IRF value increased in the AB group. The IRF percentage can be used to predict the clinical severity of AB in children.

Screening for hereditary spherocytosis in daily practice: what is the best algorithm using erythrocyte and reticulocyte parameters?

Hereditary spherocytosis (HS) is the most common inherited chronic haemolytic anaemia in Northern Europe. During the last decade, additional erythrocyte and reticulocyte parameters have been developed on last-generation haematology analysers, leading to many publications about their effectiveness as a HS screening tool. For the first time on an independent cohort, we evaluated and compared the effectiveness of six published algorithms for the screening of HS using the UniCel DxH800 (Beckman-Coulter) and the XN-9000 (Sysmex) and determined which algorithm could be the most suitable in our daily clinical practice. A total of 95 EDTA samples were analysed prospectively on both haematology analysers. These included 11 confirmed HS patients and 84 non-HS patients. The specific reticulocyte parameters used on the DxH800 were mean reticulocyte volume, immature reticulocyte fraction and mean sphered cell volume, and on the XN-9000 were hypohaemoglobinised erythrocytes, microcytic erythrocytes and immature reticulocyte fraction. The three algorithms using parameters specific to Beckman-Coulter analysers provided a sensitivity of 100% with various specificities, ranging from 7.1 to 73.8%. The three algorithms published based on the parameters specific to Sysmex showed much lower performances, i.e. out of the 11 patients with HS, between one to five patients were screened as negative for HS. However, 100% sensitivity and specificity were reached using the EMA binding test concomitantly with those three algorithms. The algorithms using reticulocyte and erythrocyte parameters offered by the recent analysers are promising options as a HS first-tier screening tool. Nevertheless, they must be evaluated by each laboratory on their own analyser before implementation.

Assessment of Reticulocyte and Erythrocyte Parameters From Automated Blood Counts in Vaso-Occlusive Crisis on Sickle Cell Disease.

Sickle cell disease is a complex genetic disease involving cell adhesion between red blood cells, white blood cells, platelets and endothelial cells, inducing painful vaso-occlusive crisis (VOC). We assessed reticulocyte and erythrocyte parameters in a cohort of confirmed SCD patients, and investigated whether a combination of these routine laboratory biomarkers of haemolysis could be used to predict VOC development. Reticulocyte and erythrocyte parameters were evaluated using the Sysmex XN-9000 analyser. A total of 98 patients with SCD were included, 72 in steady state and 26 in VOC. Among the 72 patients in steady state, 22 developed a VOC in the following year (median: 3 months [2–6]). The following parameters were increased in SCD patients with VOC development compared to SCD patients without VOC development in the following year: reticulocyte count (94.6 109/L [67.8–128] vs. 48.4 109/L [24.9–87.5]), immature reticulocyte count (259 109/L [181–334] vs. 152 109/L [129–208]) reticulocyte/immature reticulocyte fraction (IRF) ratio (6.63 109/(L*%) [4.67–9.56] vs. 4.94 109/(L*%) [3.96–6.61]), and medium fluorescence reticulocytes (MFR) (19.9% [17.4–20.7] vs. 17.1% [15.95–19.75]). The association of a reticulocyte count of >189.4 109/L and an MFR of >19.75% showed a sensitivity of 81.8% and a specificity of 88% to predict VOC development in the following year. Based on our findings, a combination of routine laboratory biomarkers, as reticulocyte count, immature reticulocyte count and fluorescent reticulocyte fraction at steady state, could be used to predict VOC development in SCD.

Hematological changes in neonatal mice with phlebotomy-induced anemia.

Anemia is a nearly universal diagnosis in preterm infants, caused by phlebotomy, and exacerbated by the underlying erythropoietic immaturity. Newborn infants are exposed to the unique stressor of fetal-to-neonatal transition, which requires significant adaptation ex utero. Accordingly, the preterm infant's response to anemia may alter the ability to confront underlying illness. This study utilized our preclinical mouse model of phlebotomy-induced anemia (PIA) to comprehensively investigate associated hematological changes. C57BL/6 mice were subjected to timed phlebotomy between postnatal days 2--10 to induce severe anemia. Complete blood counts were determined by the Sysmex XT-2000iV analyzer. Anemic pups showed a gradual reduction of RBC and hemoglobin (Hb) and increased reticulocyte (RET) counts and red cell distribution width (RDW), however, with reduced RET-Hb from postnatal day (P) of 4 onwards. Elevated levels of high fluorescent RET and immature reticulocyte fraction (IRF) were noted in anemic mouse pups, but low and medium fluorescent RET were reduced. Also, the reduction of mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), and mean corpuscular hemoglobin concentration (MCHC) were noted in anemic pups. No changes were seen in lymphocytes, but monocytes and neutrophils were significantly elevated from P4-P6. PIA in mouse pups is associated with hematological changes that may be exacerbating factors in neonatal diseases. Anemia is common and often severe in premature infants. Investigation of hematological parameters in settings of preclinical anemia may be an index of therapeutic strategies. Preclinical model evaluating the effects of neonatal anemia on the remainder of complete blood count. Detailed time kinetic phlebotomy-induced anemic mice enable us to study the impact on developmental delays in erythropoiesis and possible strategic intervention. Hematological effects of severe anemia in mice might provide insight on how best to investigate anemia in preterm infants.

Impaired Terminal Erythroid Maturation in β0-Thalassemia/HbE Patients with Different Clinical Severity.

Anemia in β-thalassemia is associated with ineffective erythropoiesis and a shortened lifespan of erythroid cells. The limited differentiation of β-thalassemic erythroblasts has been documented, but the characteristic feature of terminal erythroid maturation and its physiological relevance are not clearly described in β-thalassemias. Here, the red blood cell and reticulocyte cellular characteristics were determined in patients with β0-thalassemia/HbE in comparison to patients with iron deficiency anemia and healthy normal subjects. Severely affected β0-thalassemia/HbE patients showed the highest increase in immature reticulocytes, but the number of total erythrocytes was the lowest. Despite similar ranges of hemoglobin levels, β0-thalassemia/HbE patients had a higher number of reticulocytes and a greater proportion of immature fraction than patients with iron deficiency anemia did. In vitro CD34+ hematopoietic progenitor cells’ culture and flow cytometry analysis were conducted to investigate the erythroid maturation and mitochondrial clearance in β0-thalassemia/HbE erythroid cells as compared to normal cells. The delayed erythroid maturation and evidence of impaired mitochondria clearance were observed in β0-thalassemia/HbE cells at the terminal stage of differentiation. Additionally, increased transcript levels of genes related to erythroid mitophagy, BNIP3L and PINK1, were revealed in β0-thalassemia/HbE erythroblasts. The findings indicate that the erythroid maturation is physiologically relevant, and that the restoration of terminal maturation represents a potential therapeutic target for β-thalassemias.

The Effect of Pre-operative Autologous Blood Donation on Bone Marrow Hematopoietic Functions in Rabbits after Hepatectomy.

Pre-operative autologous blood donation (PABD) is one of the most widely distributed autologous blood donation means, which has positive effects on erythropoiesis. However, whether PABD can stimulate the bone marrow hematopoiesis after hepatectomy has not been reported. Totally 80 New Zealand rabbits were randomly divided into 4 groups that included control group, surgery group, hemodilutional autotransfusion (HA) group and PABD group. Automatic reticulocyte examination was performed to detect the content of reticulocyte and immature reticulocyte fractions (IRF). Flow cytometric analysis was employed to monitor the level of CD34+ cells and the cell cycle status. Southern blotting was conducted to determine the telomere length of CD34+ cells. The content of high fluorescence reticulocytes (HFR) and IRF was decreased at 6 h and 24 h after autotransfusion. However, the level of CD34+ cells was upregulated after PABD. Cell cycle status analysis revealed that the majority of the CD34+ cells in HA and PABD group were maintained in G0/G1 phase. The telomere length in HA and PABD group was shortened than that of the control group and surgery group. PABD could promote the bone marrow hematopoietic functions in rabbits after hepatectomy via stimulating proliferation of CD34+ cells and shortening the telomere length of CD34+ cells, but the content of HFR was not increased immediately because of the stuck of CD34+ cells in the G0/G1 phase.

Characteristics of bone marrow compensatory erythropoiesis in hereditary spherocytosis

目的揭示遗传性球形红细胞增多症（HS）骨髓红系造血的代偿特征，探究不同程度贫血对骨髓造血代偿的影响。方法收集2014年7月至2020年9月中国医学科学院血液病医院确诊的HS患者临床及实验室资料，以外周血网织红细胞绝对值作为替代参数，评估骨髓红系造血代偿能力，并对不同贫血程度HS患者红系造血代偿进行比较。结果① 302例HS患者中代偿性溶血病（代偿组）115例，轻、中、重度贫血（失代偿组）患者分别为74、90、23例。② 失代偿组血清红细胞生成素（EPO）水平与HGB呈负相关（rs＝−0.585，P<0.001）。③ HS患者的中位网织红细胞计数（ARC）0.34（0.27，0.44）×1012/L，约为正常人的4.25倍，最大ARC 0.81×1012/L，约为正常人的10倍；代偿组的中位ARC 0.29（0.22，0.38）×1012/L，约为正常人的3.63倍，失代偿组中位ARC 0.38（0.30，0.46）×1012/L，明显高于代偿组（z＝4.999，P＝0.003），达正常人的4.75倍。④代偿组的ARC与HGB呈负相关（r＝−0.177，P＝0.002）；失代偿组ARC与HGB呈正相关（rs＝0.191，P＝0.009），轻、中、重度贫血组间ARC差异无统计学意义（χ2＝4.588，P＝0.101）。⑤ 轻、中、重度贫血组的中位未成熟网织红细胞指数（IRF）分别为13.1％（9.1％，18.4％）、17.0％（13.4％，20.8％）、17.8％（14.6％，21.8％），轻度贫血组IRF小于中度及重度贫血组（Padj值均<0.05），而中度和重度贫血组间差异无统计学意义（Padj＝1.000）；轻、中、重度贫血组的中位新生网织红细胞计数分别为5.09（2.60，7.74）×1010/L、6.24（4.34，8.83）×1010/L、7.00（3.07，8.22）×1010/L，组间差异无统计学意义（χ2＝3.081，P＝0.214）。结论HS骨髓红系造血代偿随不同程度红细胞减少而增加，维持HGB在正常水平；而一经贫血发生，造血代偿即已达最大水平，不因贫血加重、EPO增加而进一步代偿增加。

Detection of testosterone microdosing in healthy females.

The ready detectability of synthetic androgens by mass spectrometry (MS)-based antidoping tests has reoriented androgen doping to using testosterone (T), which must be distinguished from its endogenous counterpart making detection of exogenous T harder. We investigated urine and serum steroid and hematological profiling individually and combined to determine the optimal detection model for T administration in women. Twelve healthy females provided six paired blood and urine samples over 2 weeks prior to treatment consisting of 12.5-mg T in a topical transdermal gel applied daily for 7 days. Paired blood and urine samples were then obtained at the end of treatment and Days 1, 2, 4, 7, and 14 days later. Compliance with treatment and sampling was high, and no adverse effects were reported. T treatment significantly increased serum and urine T, serum dihydrotestosterone (DHT), urine 5α-androstane-3α,17β-diol (5α-diol) epitestosterone (E), and urine T/E ratio with a brief window of detection (2-4days) as well as total and immature (medium and high fluorescence) reticulocytes that remained elevated over the full 14 posttreatment days. Carbon isotope ratio MS and the OFF score and Abnormal Blood Profile score (ABPS) were not discriminatory. The optimal multivariate model to identify T exposure combined serum T, urine T/E ratio with three hematological variables (% high fluorescence reticulocytes, mean corpuscular hemoglobin, and volume) with the five variables providing 93% correct classification (4% false positive, 10% false negatives). Hence, combining select serum and urine steroid MS variables with reticulocyte measures can achieve a high but imperfect detection of T administration to healthy females.

Utility of reticulocyte indices in the diagnosis of pancytopenia.

Introduction:Pancytopenia is a hematological condition in which there is a reduction in all three cell lines of blood. This study aims to evaluate the utility of reticulocyte indices such as reticulocyte % (retic %), immature reticulocyte fraction (IRF), and mean reticulocyte volume (MRV) in identifying the cause of pancytopenia.Materials and Methods:Reticulocyte indices were measured by an automated coulter. These values were then correlated with relevant biochemical and bone marrow results and cases were stratified into different etiological groups. Receiver operator curve (ROC) analysis was performed and various cut-off values were derived based on the reticulocyte indices. ROC was repeated to further classify cut-off values at every level to help formulate a diagnostic algorithm.Results:A total of 154 cases of pancytopenia were obtained. Ages ranged from 7 months to 87 years with a mean of 42, the male:female ratio was 1.08:1. The majority of the cases were megaloblastic anemias in which the cut-off values for retic % was <0.91 with a sensitivity of 78.1% and specificity of 70%, IRF was 0.45 with a sensitivity of 76.7% and specificity of 64%, and MRV was >121.8 fl with a sensitivity of 83.6% and specificity of 80%. The values on ROC could segregate nutritional from nonnutritional causes. The IRF and MRV also helped to differentiate megaloblastic anemia from dual deficiency anemia.Conclusion:Reticulocyte indices help identify the cause of pancytopenia. They can segregate nutritional anemia from other causes of pancytopenia allowing presumptive treatment to be initiated and may obviate invasive procedures such as bone marrow examination.

Identification of subclinical iron deficiency using new erythrocytes, leukocytes, and reticulocytes parameters during nonsevere acute infection in pediatric outpatients.

This study aims to investigate the diagnostic utility of new erythrocytes, leukocytes, and reticulocytes parameters for the identification of subclinical iron deficiency (ID) in children under 6 years with nonsevere acute infection in pediatric outpatients. The study included 102 children with acute infections and 31 true ID. Traditional and new hematology parameters were measured in a Sysmex-XN®, along with C-reactive protein level, and iron parameters. Participants' ID were categorized as: the ferritin < 100 ng/mL, transferrin saturation < 20% was defined as "subclinical or functional ID (FID) in Group 1"; ferritin < 30 ng/mL, transferrin saturation < 20%, as "absolute-ID (AID)" in Group 2; ferritin < 12 ng/mL without anemia and infection, as "true ID" in Group 3. The frequencies of FID and AID among the 102 children with acute infection were 24% and 76%, respectively. Compared with the Group 2 patients, Group 1 had a significantly higher mean percentage of hypochromic erythrocytes (Hypo-He), and significantly lower levels of hemoglobin (Hb) and Hb content of reticulocytes (RET-He) (p < 0.05 for all). Compared with Group 2 and Group 3 patients, Group 1 had a significantly higher mean percentage of immature reticulocyte fraction (IRF) and immature granulocyte (IG) values (p < 0.05 for all). The RET-He, IRF%, Hypo-He%, and IG% cut-off values for predicting FID during infection were 27.0 pg, 10.6%, 2.5%, and 0.35% respectively. The RET-He, Hypo-He, IRF, and IG may be useful parameters for identifying subclinical ID in small children with nonsevere acute infection in pediatric outpatients.

Manual immature reticulocyte fraction; a surrogate marker to assess post traumatic blood loss

Introduction: Erythropoietic response following chemotherapy and stem cell transplant is assessed by reticulocytosis and immature reticulocyte fraction (IRF). They are not used to assess erythropoiesis following acute haemorrhage even though it is known to cause reticulocytosis. This study was performed to evaluate the use of IRF as a marker of degree of post-traumatic haemorrhage. Methods: A cross sectional descriptive study was performed using routine anticoagulated blood samples collected from patients admitted with acute traumatic haemorrhage to an emergency treatment unit. Differential and total reticulocyte counts were performed manually using standard techniques with bright field microscopy. The data on severity of blood loss were extracted from clinical records. Results: There were total of 38 patients. Mean IRF of patients with mild (n = 14), moderate (n = 19) and severe (n = 05) haemorrhage were 1.86 ± 1.0%, 3.26 ± 1.8% and 10.2 ± 3.6% respectively. Mean manual IRF of group with severe haemorrhage was significantly higher than those of all the others (p = 0.000). The mean manual IRF of moderate haemorrhage group was significantly higher than that of mild group (p = 0.01). Manual IRF and severity of haemorrhage were significantly positively correlated (r = 0.71, p &lt; 0.01, n = 38). Immature reticulocytes of stage I was observed only in patients with severe haemorrhage. Conclusion: The manual IRF can be used as a reliable marker of severity of post-traumatic acute haemorrhage.

Reference intervals for reticulocyte parameters (reticulocyte haemoglobin equivalent and immature reticulocyte fraction) in first-trimester pregnancy

Reticulocyte parameters including reticulocyte haemoglobin equivalent (Ret-He) and immature reticulocyte fraction (IRF) are newly recognised hematological parameters that are being used for diagnosis and follow-up of anaemic patients. Reference intervals of these parameters have been established in different populations, however, the data relating to pregnancy are still lacking. One hundred and fifty-five first-trimester pregnant females were screened and the reference interval was calculated after selecting the patient with fixed criteria. R statistical software was used for statistical calculations. We tried to establish a reference interval of Ret-He content and IRF in first-trimester pregnancy in our study. IMPACT STATEMENT What is already known on this subject? Ret-He and IRF have been established as the marker of iron deficiency and iron-deficiency anaemia in different age groups and as a marker of response to iron therapy. However, literature is scarce regarding the reference intervals of these parameters, especially in pregnancy. What do the results of this study add? This study establishes the reference interval of newer reticulocyte parameters in first-trimester pregnancy which is not yet established in the literature. Establishing a reference interval is required for any laboratory parameters to be used in the clinical context. What are the implications of these findings for clinical practice and further research? The results of this study may help in making a clinical decision regarding iron deficiency in early pregnancy which is one of the common clinical problems in pregnancy. This study also serves as a baseline study for further studies of reference intervals for newer reticulocyte parameters in pregnancy. A similar study with a larger study population and follow-up with iron therapy may establish these parameters as one of the important markers of iron deficiency in pregnancy and help institute iron therapy on case-to-case basis.

Reference intervals of automated reticulocyte count and immature reticulocyte fraction in a pediatric population.

Reticulocytes are erythroid precursors that develop into mature erythrocytes, and they are an important tool to assess erythropoietic activity, as their count indicates the balance between the cells released from the bone marrow, their stage of maturity, and their rate of development into mature erythrocytes. Considering the described biological variability of the absolute reticulocyte count (ARC) and the immature reticulocyte fraction (IRF) and the limited information available on these hematological parameters in children, this study determined the reference intervals (RIs) of these parameters in a healthy pediatric population. A retrospective, observational, and analytical study was designed to establish RIs for the ARC and the IRF according to age and sex. An indirect sampling method was applied to a mixed database of complete blood counts from children aged 2months to 18years, using the truncated maximum likelihood indirect method for reference interval estimation. Percentiles were calculated to obtain bimodal RIs. From a total of 190,812samples, 6,814 were selected. Gender stratification was not necessary for the ARC and the IRF but they required partitioning into six and two age groups, respectively. This study determined, by an indirect sampling method, RIs for the ARC and the IRF in a pediatric population according to age and sex.

Difference in Immature Reticulocyte Fraction Percentage between Moderate and Severe Anemia in Transfusion-Dependent Thalassemia

Thalassemia is an inherited genetic disease caused by the disruption in globin chain synthesis. Inefective erythropoiesis in thalassemia leads to moderate to severe anemia, requiring routine blood transfusions. To evaluate erythropoiesis, immature reticulocyte fractions (IRF) can be measured using the hematology analyzer, avoiding the need of invasive bone marrow examination. The purpose of this study was to analyze the differences in the IRF percentage between moderate and severe anemia in transfusion-dependent thalassemia (TDT) patients. This was a cross-sectional comparative observational analytic study conducted at the Pediatric Thalassemia Clinic of Dr. Hasan Sadikin General Hospital Bandung in August–September 2020. The IRF was examined using the fluorescence flowcytometry method with whole blood sample added by EDTA anticoagulant. The statistical analysis used in this study was unpaired t-test and Mann Whitney’s test. Subjects were 93 TDT pediatric patients, consisting of 48 boys (52%) and 45 girls (48%). The majority (72%) of the patients had been diagnosed with thalassemia for more than 5 years with moderate anemia (40%) and severe anemia (60%). The median IRF percentage in moderate anemia was 6.4% (range 0-22.7) while the range in severe anemia was 11.7% (range 4.1–35.8), suggesting a statistically significant difference (p&lt;0.001) in the IRF percentage between moderate and severe anemia in transfusion-dependent thalassemia patients. To conclude, the more severe the anemia experienced by a thalassemia patient is, the higher the percentage of IRF.

Utility of reticulocyte haemoglobin content and immature reticulocyte fraction in early diagnosis of latent iron deficiency in whole blood donors.

The present study was planned to assess the clinical utility of reticulocyte haemoglobin content (CHr) and immature reticulocyte fraction (IRF) in the early detection of latent iron deficiency in blood donors. The prospective longitudinal observational study was conducted using the purposive sampling method. Written informed consent was obtained and donors were allocated into the first-time (FTD) and regular donor (RD) group. The enrolled blood donors (n= 205 in each group) were followed up for two subsequent whole blood donations. Haemoglobin (Hb), CHr, IRF and serum ferritin values were recorded at enrolment and two follow-ups. The sensitivity of CHr in detecting iron-deficient erythropoiesis (serum ferritin values ≤ 26 μg/dl) was 45% and 56.7%, specificity 96.7%, positive predictive value (PPV) 85.6% and 90.8% and negative predictive value (NPV) 80.1% and 78.7%, respectively in FTD and RD cohorts. The sensitivity of IRF was 45.1% and 44.8%, specificity 93.4% and 97.1%, PPV 74.8% and 90.4% and NPV 79.6% and 74.5%, respectively in both the cohorts. The sensitivity of CHr in detecting absent iron stores (serum ferritin values ≤ 15 μg/dl) was 66.2% and 74.4%, specificity 92% and 90.6%, PPV 56.7% and 68.7% and NPV 94.5% and 92.8% among FTD and RD cohort, respectively. The sensitivity of IRF was 72.7% and 65.3%, specificity 90.3% and 94.3%, PPV 54.4% and 76% and NPV 95.4% and 90.8%, respectively in both the cohorts. Reticulocyte hemoglobin content and IRF can be used along with complete blood count for early detection of iron deficiency in blood donors using the same blood sample at no extra cost.

Etavopivat, an Allosteric Activator of Pyruvate Kinase-R, Improves Sickle RBC Functional Health and Survival and Reduces Systemic Markers of Inflammation and Hypercoagulability in Patients with Sickle Cell Disease: An Analysis of Exploratory Studies in a Phase 1 Study

Etavopivat, an Allosteric Activator of Pyruvate Kinase-R, Improves Sickle RBC Functional Health and Survival and Reduces Systemic Markers of Inflammation and Hypercoagulability in Patients with Sickle Cell Disease: An Analysis of Exploratory Studies in a Phase 1 Study

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

Abstract 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.

Tracking immature reticulocyte proteins for improved detection of recombinant human erythropoietin (rhEPO) abuse.

Athletes abuse recombinant human erythropoietin (rhEPO) and erythropoiesis stimulating agents to increase hemoglobin mass and improve performance. To evade detection, athletes have developed sophisticated blood doping regimens, which often include rhEPO micro-dosing. Detection of these methods requires biomarkers with increased sensitivity and a sample matrix that is more amenable to frequent testing in the field. We have developed a method to measure two immature reticulocyte proteins, CD71 and ferrochelatase (FECH), and one total erythrocyte protein, Band 3, in dried blood spots (DBS). This method was tested in response to rhEPO administration after low doses, 40 IU/kg, micro-doses, 900 IU, or saline injection in 20 healthy subjects. During administration of low-dose rhEPO, the mean CD71/Band 3 and FECH/Band 3 ratio increased by 412 ± 197% and 250 ± 44%, respectively. The mean response for the current biomarker, RET%, increased by 195 ± 35%. During administration of rhEPO micro-doses, CD71/Band 3 increased to 127 ± 25% on day 35 and 139 ± 36% on day 39, while no increase was observed in RET%. After rhEPO administration, during the washout phase, mean values decreased to a minimum of 64 ± 4% and 64 ± 11% for CD71/Band 3 and RET%, respectively. However, CD71/Band 3 remained below 75% of baseline for at least 4 weeks after rhEPO injection, while RET% returned to baseline levels. The results demonstrate that immature reticulocyte proteins have a larger response to rhEPO administration than the current biomarker, RET%, and can be monitored in the DBS matrix.

Comparison of the value of immature retyculocyte and immature platelet in the diagnosıs of sepsis.

Sepsis is one of the causes of pre-treatment morbidity and mortality in the pediatric age group. In the present study, we investigated the place of the immature granulocyte percentage, (IG) immature reticulocyte fraction (IRF), and immature platelet fraction (IPF) in the diagnosis of sepsis. Complete blood count, C-reactive protein, (CRP) procalcitonin (PCT) and blood cultures were measured in 125 critical patients who were followed-up in the intensive care unit with the suspicion of sepsis and 65 healthy children between 2017 and 2019. In addition to the complete blood counts and routine parameters, IG, IRF, and IPF were examined in the patients. When the critical patient group and the healthy control group were compared, it was found that the total number of leukocytes (white blood cells), neutrophil count, platelet count, CRP, PCT, IG, IRF, and IPF values were higher at statistically significant levels. When septic and non-septic patients were compared, it was found that the CRP, PCT,IGP, and IPF were higher at statistically significant levels in the septic patients. It was concluded that CRP, PCT, IG, and IPF were significant in determining sepsis and that PCT was the most sensitive and specific biomarker in these parameters. We believe that these parameters may be suitable for practical use in determining sepsis because they give faster results and suggest the diagnosis of sepsis.

Plasmodium vivax binds host CD98hc (SLC3A2) to enter immature red blood cells.

More than one-third of the world's population is exposed to Plasmodium vivax malaria, mainly in Asia1. P. vivax preferentially invades reticulocytes (immature red blood cells)2-4. Previous work has identified 11 parasite proteins involved in reticulocyte invasion, including erythrocyte binding protein 2 (ref. 5) and the reticulocyte-binding proteins (PvRBPs)6-10. PvRBP2b binds to the transferrin receptor CD71 (ref. 11), which is selectively expressed on immature reticulocytes12. Here, we identified CD98 heavy chain (CD98), a heteromeric amino acid transporter from the SLC3 family (also known as SLCA2), as a reticulocyte-specific receptor for the PvRBP2a parasite ligand using mass spectrometry, flow cytometry, biochemical and parasite invasion assays. We characterized the expression level of CD98 at the surface of immature reticulocytes (CD71+) and identified an interaction between CD98 and PvRBP2a expressed at the merozoite surface. Our results identify CD98 as an additional host membrane protein, besides CD71, that is directly associated with P. vivax reticulocyte tropism. These findings highlight the potential of using PvRBP2a as a vaccine target against P. vivax malaria.