OBJECTIVE: Women are more prone to iron deficiency (ID) anemia when pregnant. The diagnostic use of mean reticulocyte volume (MRV) in identifying ID anemia during pregnancy has not been thoroughly investigated. The objective of this study is to evaluate the effectiveness of MRV in diagnosing ID in pregnant women.
METHODS: Firstly, MRV of 20 healthy female volunteers (healthy group) was measured on specific days for one month. Subsequently, clinical data from 724 pregnant women were thoroughly examined. These women were divided into two groups: 282 with ID (research group) and 442 without ID (control group). Parameters such as MRV, reticulocyte hemoglobin equivalent (RHE), red blood cell volume distribution width-standard deviation (RDW-SD), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), hematocrit (HCT), reticulocyte count (RET), MRV/MCV ratio, and serum ferritin (SF) were analyzed and compared.
RESULTS: MRV remained consistent over a period of one month for 20 healthy individuals. In addition, there were significant differences in MRV, RHE, RDW-SD, MCV, MCH, MCHC, HCT, RET, and MRV/MCV between the research group and control group. The receiver operating characteristic (ROC) analysis showed that the areas under the curve (AUCs) for these measures were as follow: 0.840, 0.837, 0.676, 0.654, 0.639, 0.602, 0.571, 0.550, and 0.816, respectively. Ultimately, there was a substantial disparity in MRV prior to and following therapy with oral iron treatments.
CONCLUSIONS: In healthy women, MRV remains stable and is a reliable ID marker, which can be used to assess oral iron treatment effectiveness during pregnancy.

OBJECTIVE: This first-in-human trial aimed to investigate the pharmacokinetics and pharmacodynamics characteristics and safety and tolerability of single ascending doses of subcutaneous polyethylene glycol-erythropoietin (PEG-EPO) in healthy subjects.
METHODS: In this phase I, randomized, double-blind, placebo-controlled, dose-escalating trial, subjects were sequentially enrolled into 7 cohorts with 12 subjects in each cohort and randomized in a 5:1 ratio to receive a single dose of 0.2, 0.4, 0.8, 1.6, 2.4, 3.6, or 4.8 µg/kg PEG-EPO or matching placebo. Safety and tolerability including dose-limiting toxicities (DLTs) were assessed. Pharmacokinetics parameters, including Cmax, AUC0-inf, Tmax, and t1/2, and pharmacodynamics parameters, including reticulocyte count and hemoglobin content, were evaluated.
RESULTS: Eighty-four subjects (median age 30.4 years and 77.4% male) were enrolled. No subjects developed DLTs. Any grade treatment-related adverse events occurred in 66.7% of the subjects, but most (92.9%) were mild. No serious adverse events and no death occurred. Forty percent of the subjects receiving PEG-EPO had iron decreased, 27.1% reported ferritin decreased, 25.7% showed unsaturated iron binding capacity increased, and 17.1% had neutrophil count decreased. Cmax exhibited a dose-disproportionate rise from a geometric mean of 525 pg/mL with 0.2 µg/kg PEG-EPO to 23196 pg/mL with 4.8 µg/kg PEG-EPO. The mean t1/2 ranged between 82.4 ± 21.3 h with 0.4 µg/kg PEG-EPO and 160.6 ± 65.7 h with 1.6 µg/kg PEG-EPO. AUC0-inf displayed a largely dose-proportional rise from 226264.5 pg*h/mL with 0.2 µg/kg PEG-EPO to 5206434.0 pg*h/mL with 4.8 µg/kg PEG-EPO. The absolute reticulocyte count increased with escalating doses of PEG-EPO, with the mean maximal change from baseline between 3.2 ± 1.5*10^10/L (Q1,Q3 1.8-3.6*10^10/L) with PEG-EPO 0.2 µg/kg and 9.3 ± 4.0*10^10/L (Q1,Q3 6.2-13.5*10^10/L) with 3.6 µg/kg PEG-EPO. The mean maximal change from baseline in the mean hemoglobin content ranged between 5.9 ± 4.4 g/L (Q1,Q3 3.5,7.0) with 0.2 µg/kg PEG-EPO and 15.4 ± 8.7 g/L (Q1,Q3 10.5,20.0) with 2.4 µg/kg PEG-EPO.
CONCLUSIONS: This trial demonstrated that PEG-EPO was safe and tolerable in healthy subjects. The subcutaneous route of administration allows outpatient treatment and the pharmacokinetics characteristics of PEG-EPO support less frequent dosing regimens and effective treatment for chronic kidney disease patients with anemia.
BACKGROUND: clinicaltrials.gov identifier: NCT03657238.

OBJECTIVE: To determine, among neonates at-risk for hyperbilirubinemia, whether measuring end-tidal carbon monoxide concentration (ETCOc) twice before 48 hours could identify those who would develop hyperbilirubinemia and differentiate hemolytic vs. non-hemolytic causes.
METHODS: Prospective study on neonates meeting criteria \"at-risk for hyperbilirubinemia.\" Routine bilirubin measurements and 10-day follow-up were used to categorize neonates as; (1) normal (no hyperbilirubinemia, all bilirubins <95th percentile of Bhutani nomogram), (2) having hemolytic hyperbilirubinemia (bilirubin ≥95th percentile, DAT+, elevated retic, or G6PD+), or (3) having non-hemolytic hyperbilirubinemia.
RESULTS: 386 neonates were enrolled. 321 (83%) did not develop hyperbilirubinemia and 65 (17%) did, of which 29 were judged hemolytic and 36 non-hemolytic. High ETCOc differentiated the hemolytic group (p < 0.001). First-day ETCOc correlated with bilirubin and with reticulocyte count (r = 0.896 and 0.878) and sensitivity and specificity for predicting hyperbilirubinemia were excellent (83% and 95%).
CONCLUSIONS: ETCO measurement in the first 48 hours after birth predicts hemolytic hyperbilirubinemia.

BACKGROUND: The Sysmex XN-1000V automated hematology analyzer with multispecies software was released in June 2017 for use in research laboratories. Laser light, impedance, fluorescent staining, and fluorescent flow cytometry are used to analyze whole blood for CBC, reticulocyte counts, and WBC counts, including a 5-part differential leukocyte analysis.
OBJECTIVE: A side-by-side comparison of the Sysmex XN-1000V with the Siemens ADVIA 120 in analyzing blood from healthy mice and rats will provide insight into the performance of the new analyzer and its capabilities for use in drug development studies. Method correlation analyses on normal mouse and rat hematology data collected with both analyzers and manual reference methods will help determine the reliability of the data produced using the Sysmex XN-1000V analyzer.
METHODS: Whole blood samples collected in K2 EDTA from healthy CD-1 mice and CD Sprague-Dawley rats were analyzed in parallel with the XN-1000V and ADVIA 120 analyzers. Male and female mice, approximately 6-9 weeks old, and male and female rats, approximately 7-9 weeks old, were included in this study. Manual reference methods for WBC differential leukocyte analysis and packed cell volume (PCV) measurements were also performed. EP Evaluator version 11.2 (Data Innovations LLC, South Burlington, VT, USA) was used for method comparison statistical analysis.
RESULTS: Most hematologic parameters for naïve mice and rats achieved correlation in the fair to excellent range, with the majority showing very good to excellent correlation with low biases (<11.0%) for cohorts analyzed separately and when cohort data were combined.
CONCLUSIONS: The Sysmex XN-1000V Hematology Analyzer provided comparable results to those obtained from the Siemens ADVIA 120. We found the Sysmex XN-1000V Hematology Analyzer to be acceptable for use in drug development studies for rats and mice.

BACKGROUND: In the modern era of complete blood count analysis, manual differential count is performed whenever \'flags\' are generated by an automated hematology analyzer. Traditionally, tally counters with five or eight keys are used for manual differential count. A few mobile applications are available to perform this task; however, the application features and cell representation are limited.
OBJECTIVE: The primary objective of our study was to develop an indigenous, comprehensive mobile application to assist with manual blood cell differential count. The secondary objective was to measure the usability of a newly developed application among undergraduate medical students.
METHODS: A new mobile application was developed using a Java development kit, Version 11.0.13 (Oracle Corporation, Austin, USA) in Android Studio Dolphin (2021.3.1) (Google, California, USA). The application content was validated by three pathologists with more than five years of experience. The app\'s usability was tested among 60 participants using a validated mHealth App Usability Questionnaire (MAUQ). The questionnaire had 18 items covering three domains: ease of use, interface & satisfaction, and usefulness.
RESULTS: The newly developed application supports peripheral smear WBC differential count, platelet count, reticulocyte count, malaria parasite quantification, and bone marrow differential count. During usability testing, the app was easy to use in 95% (57/60) of participants, time-efficient in 91.7% (55/60), and helpful for healthcare practice learning in 96.7% (58/60). The total mean score was 6.11, indicating high usability.
CONCLUSIONS: A comprehensive mobile application to assist manual differential count with adequate cell representation was developed. The mobile application was easy to use, time-efficient, and valuable among the study participants.

BACKGROUND: The International Council for Standardization in Haematology convened a working group to assess and propose improvements upon the state of standardization and harmonization of reticulocyte parameters among commercial hematology analyzers.
METHODS: An international group of laboratory hematologists prospectively collected and analyzed clinical samples using locally available IVD commercial hematology analyzers. Eight hundred and fifty-five total samples were collected at 6 sites using 9 distinct analyzer types. Samples were assessed for reticulocyte percent (RET%), immature reticulocyte fraction (IRF), and reticulocyte hemoglobin content (RHC). Method comparison and regression statistics were calculated. These analyses were used to determine whether statistical recalibration offered a potential avenue for increasing comparability between these methods.
RESULTS: While methods producing reticulocyte percent were the most comparable in this study, the state of harmonization for the IRF and RHC was reduced with pearson correlation coefficients ranging from 0.955 to 0.77 and 0.927 and 0.680, respectively. Nevertheless, use of parameters from the Passing Bablok regression substantially improved the comparability of the results. In addition, precision data was derived which also demonstrated substantial differences between analyzer systems.
CONCLUSIONS: While reticulocyte counting is correlated between the automated methods evaluated in this study, the current state of harmonization of other reticulocyte parameters is not as strong. A major challenge in moving this field forward is the need for commutable materials to facilitate comparisons between analyzers not co-located. A potential alternate approach to improve the current state would be instrument re-calibration. However, this is challenging both technically and due to national regulatory frameworks.

Transcatheter aortic valve implantation (TAVI) has brought in recent years relief of cardiac-induced symptoms to a large number of patients with aortic stenosis. Whether it is better to use TAVI for the treatment of aortic valve stenosis superimposed on a congenitally bicuspid valve has been debated in contrast to its proved usefulness in aortic valve stenosis involving a tricuspid aortic valve. From January 2020 to March 2023, surgical aortic valve replacement of TAVI valve and native aortic valve was done in 6 patients. The clinical findings of the patients and morphologic findings from the surgical specimens submitted to the cardiac pathology department were subsequently examined. All the 6 native aortic valves had bicuspid configuration. The TAVI valve in each patient was excised from 9 to 88 months (mean 36 months) after it had been implanted because of paravalvular leak in 4, severe stenosis of the prosthetic valve in 1, and bioprosthetic cuspal degeneration in 1. Prosthetic valve endocarditis was clinically suspected in 2 patients, but the specimen culture was negative. Before surgical aortic valve replacement, 3 patients experienced stroke after TAVI. All 6 patients had low hemoglobin levels (mean 9.5 mg/100 ml) and low hematocrit levels (mean 29.5%). Reticulocyte count was available in 4 patients and was increased in all (mean 3.5%). When the stenotic native aortic valve configuration is bicuspid, the raphe tends to be calcified first and located perpendicular to the flow of the blood and may prevent the ring of the caged bioprosthesis from being transferred to the aortic wall, which is a requirement for full opening of the lumen of the bioprosthesis. Thus, thorough consideration needs to be made before performing TAVI in patients whose native aortic valve is stenotic and bicuspid.

Reticulocyte indices are used to characterize anemia, including the identification of regeneration. In people, the immature reticulocyte fraction (IRF), percentage of hypochromic red blood cells (%HYPO-RBC), and other reticulocyte indices have been used as earlier indicators of erythropoiesis and as valuable monitoring tools in the assessment of various therapies. The reference intervals (RI) of the IRF and %HYPO-RBC have not been reported in dogs. The objective of this study was to establish RIs for novel variables (IRF, %HYPO-RBC, and CH-delta) and assess RIs for more commonly reported reticulocyte indices in healthy dogs. RIs were calculated from blood results retrospectively collected from 106 client-owned healthy dogs at the time of induction into a blood donor program using the ADVIA 2120 hematology analyzer (Siemens Healthcare Diagnostics). For the calculation of RIs, appropriate tests were applied for outlier detection and normality assessment. For variables normally distributed, RIs and their respective 90% confidence intervals (CIs) were calculated using parametric methods, while for variables not normally distributed, robust methods were used and bootstrapping for calculating the 90% CIs. The following RIs were established: reticulocyte hemoglobin content (CHr) 24.5-28 pg, mean reticulocyte volume (MCVr) 85.9-99.3 fL, mean corpuscular hemoglobin concentration of reticulocytes (CHCMr) 271.0-306.3 g/L, IRF 10.4%-43.5%, CH-delta 0.5-4.3 pg, and percentage of hypochromic red blood cells (%HYPO-RBC) 0.10%-0.80%. The results of this study provide RIs for novel reticulocyte variables. Further studies are required to determine the clinical utility of IRF, %HYPO-RBC, and CH delta as early indicators of erythropoietic activity in canine patients.

OBJECTIVE: We investigated the potential of reticulocyte haemoglobin equivalent (RET-He) as an early marker of responsiveness to iron supplementation.
METHODS: Data were obtained from a randomised controlled trial of daily iron supplementation in 356 Cambodian women (18-45 y) who received 60 mg elemental iron for 12 weeks. A fasted venous blood specimen was collected at baseline, 1-week and 12-week timepoints. Whole blood haemoglobin (g/L) and RET-He (pg) were measured using a Sysmex haematology analyser. RET-He measures were evaluated for their predictive ability on haemoglobin response to iron supplementation (defined as ≥10 g/L at 12 weeks). Receiver operating characteristic (ROC) curves were used to assess discrimination performance, and the area under the ROC curve (AUCROC) served as a measure of the ability of each predictor to discriminate between women likely or unlikely to elicit a haemoglobin response.
RESULTS: Predictive ability (AUCROC (95% CI)) of baseline, 1-week, and change from baseline to 1-week RET-He on haemoglobin response was 0.70 (0.63 to 0.76), 0.48 (0.41 to 0.56) and 0.81 (0.75 to 0.87), respectively. Based on the Youden index, an absolute increase in RET-He of ~1.1 pg or a percentage increase of ~4.4% over 1 week were optimal thresholds to predict responsiveness to iron supplementation.
CONCLUSIONS: Single timepoint RET-He measures have poor predictive ability; however, change in RET-He after 1 week was a strong predictor of haemoglobin response among Cambodian women receiving 60 mg elemental iron and can be measured easily and quickly after only 1 week of iron therapy.

Automated fluorescence-based haematology analysers are now available for reticulocyte enumeration in veterinary medicine, but manual counting is still largely used. This study aimed to evaluate potential sources of analytical and pre-analytical errors when performing automated and manual counts.
Automated and two-operator double-blind manual reticulocyte counts were performed on 15 blood samples. The intra-assay variation of the automated and manual counts and the interoperator variation in the manual counts were then calculated. In addition, the effects of storage were evaluated using samples refrigerated at 4°C or stored at room temperature for 2, 4, 12, 24, 48 or 72 hours after sampling.
Intra-assay coefficients of variation were lower for automated counts than for manual counts. Comparison between automated and mean total manual reticulocyte count showed no significant differences. In both refrigerated samples and those stored at room temperature, an increase in reticulocyte count was recorded only after 72 hours. Staining artefacts occurred only in one stored sample counted manually.
The presence of cytoplasmic particles other than RNA can cause misinterpretation of cells, leading to an erroneous reticulocyte count.
The use of an automated analyser is preferable for reticulocyte enumeration in dogs. Common storage conditions seem to minimally affect reticulocyte evaluation; however, it is recommended to perform the analysis as soon as possible after sampling.