BACKGROUND: BCR-ABL1-negative myeloproliferative neoplasms (MPNs) often exhibit overlapping clinical and morphological features, making accurate subcategorization challenging. The h-MICL (CD371) antigen, selectively expr...BACKGROUND: BCR-ABL1-negative myeloproliferative neoplasms (MPNs) often exhibit overlapping clinical and morphological features, making accurate subcategorization challenging. The h-MICL (CD371) antigen, selectively expressed on leukemic stem cells (LSCs) and absent on CD34CD38 cells in normal or regenerating marrow, represents a potential diagnostic and prognostic marker. OBJECTIVES: This study aimed to evaluate the diagnostic utility of circulating CD34CD38h-MICL cells in subtyping BCR-ABL1-negative MPNs and to assess their correlation with the Dynamic International Prognostic Scoring System (DIPSS) score. Additionally, to identify a cut-off value of CD34CD38h-MICL cells that can effectively differentiate PMF. METHODS: Fifty-four patients with BCR-ABL1-negative MPNs were prospectively enrolled at a tertiary care center in North India over 18 months. Peripheral blood was analyzed via flow cytometry to quantify CD34, CD34CD38, CD34CD38, and CD34CD38h-MICL cell subsets. RESULTS: A significant increase in circulating CD34CD38h-MICL cells was observed in patients with overt MF (median 2.8%), prefibrotic MF (4.15%), and post-PV/ET MF (3%) compared to PV and ET (median 0%; p < 0.001). A threshold of ≥ 0.9% CD34CD38h-MICL cells effectively identified MF cases with 88% sensitivity and 89% specificity. Moreover, a positive correlation was found between the percentage of CD34CD38h-MICL cells and DIPSS score (ρ = 0.41, p = 0.036), with every 1.72% increase in this cell population corresponding to a 1-point rise in DIPSS score. CONCLUSION: Circulating CD34CD38h-MICL cells represent a promising biomarker; their quantification may aid in subclassification, particularly in distinguishing prefibrotic MF from ET, and may serve as a potential target for future therapeutic strategies. Further validation in larger cohorts is warranted.
INTRODUCTION: Fibrinogen deficiency is common in acute haemorrhage and requires prompt treatment to prevent exacerbation of coagulopathy. Conventional lab measurements are time consuming, leading to delayed intervention...INTRODUCTION: Fibrinogen deficiency is common in acute haemorrhage and requires prompt treatment to prevent exacerbation of coagulopathy. Conventional lab measurements are time consuming, leading to delayed intervention or empirical fibrinogen replacement, potentially exposing patients to unnecessary transfusion. The qLabs FIB system offers rapid point-of-care measurement of fibrinogen at the bedside; however, its use in patients taking Direct Oral Anticoagulants (DOACs) has not been evaluated. METHODS: Thirty patients chronically anticoagulated with Apixaban or Edoxaban undergoing catheter ablation procedures for atrial fibrillation were recruited. Citrated blood samples were taken from each patient and assessed using the qLab FIB system, and lab-based Clauss assay (HemosIL Q.F.A. Thrombin (QFA), Werfen, Bedford, USA). DOAC levels were also measured, then removed using DOAC-Remove (Quadratech, UK) to assess their impact. Statistical analysis was performed using Bland-Altman, Passing-Bablok regression and paired t-tests. RESULTS: A strong linear correlation (r = 0.88) was observed between the two methods; however, the qLabs FIB system consistently overestimated fibrinogen levels compared to the Clauss assay, with a mean bias of +0.50 g/L (95% LOA: -0.03 to +1.02 g/L). The Passing-Bablok slope of 0.64 suggested that overestimation might be more pronounced at lower fibrinogen concentrations. DOAC presence did not significantly alter measurement results (p = 0.380), and DOAC concentration did not correlate with the measurement discrepancy. CONCLUSION: The qLabs FIB system is a viable method for rapid assessment of fibrinogen levels in patients anticoagulated with DOACs. However, the presence of overestimation bias may have implications for clinical decision making.
INTRODUCTION: Rapid immunoassays are widely used for diagnosing heparin-induced thrombocytopenia (HIT). The STic Expert HIT kit is commonly employed, but its diagnostic accuracy in unselected, real-world populations requ...INTRODUCTION: Rapid immunoassays are widely used for diagnosing heparin-induced thrombocytopenia (HIT). The STic Expert HIT kit is commonly employed, but its diagnostic accuracy in unselected, real-world populations requires robust evaluation against an appropriate reference standard. This study evaluates the diagnostic accuracy of the STic Expert HIT kit using the heparin-induced platelet aggregation test (PAT) as the comparator. METHODS: This cross-sectional diagnostic accuracy study was conducted from January 2023 to April 2025. A consecutive series of 320 patients with suspected HIT underwent parallel testing with the STic Expert HIT kit (index test) and PAT (comparator). The 4T score was calculated prospectively for all patients. Borderline visual results on the STic kit were classified according to epidemiological recommendations: as false negative if PAT-positive and false positive if PAT-negative. Diagnostic accuracy measures-sensitivity, specificity, predictive values, and likelihood ratios-were calculated with 95% confidence intervals. The study is reported in accordance with the STARD 2015 checklist. RESULTS: Among 320 consecutive patients (median age 45.6 years; range 0.19-89), the prevalence of PAT-confirmed HIT was 16.3% (52/320). Following the STARD guidelines, the STic Expert HIT kit demonstrated a sensitivity of 57.5% (95% CI: 42.2-71.6) and specificity of 92.5% (95% CI: 88.8-95.1). The positive predictive value was 58.8% (95% CI: 46.2-70.4) and negative predictive value was 92.1% (95% CI: 89.6-94.1). The positive likelihood ratio was 7.67 (95% CI: 5.10-11.52) and negative likelihood ratio was 0.46 (95% CI: 0.32-0.66). The area under the ROC curve was 0.75 (95% CI: 0.67-0.83). Borderline results constituted 6.6% (21/320) of tests; of these, 33.3% (7/21) were true positives and 66.7% (14/21) were false positives. CONCLUSION: In this large, real-world diagnostic accuracy study, the STic Expert HIT kit demonstrated limited sensitivity for HIT diagnosis. The test is not suitable for ruling out HIT due to its suboptimal sensitivity. Borderline results are common and require integration with pre-test probability. These findings highlight the need for confirmatory functional testing in suspected HIT, particularly in high-probability settings.
INTRODUCTION: Pyruvate kinase deficiency (PKD) is a rare cause of hereditary non-spherocytic hemolytic anemia. This study reports the clinical and molecular characterization of three pediatric PKD cases, focusing on two...INTRODUCTION: Pyruvate kinase deficiency (PKD) is a rare cause of hereditary non-spherocytic hemolytic anemia. This study reports the clinical and molecular characterization of three pediatric PKD cases, focusing on two novel PKLR variants. METHODS: Clinical and laboratory data were reviewed. PKLR gene was analyzed by next-generation sequencing and validated by Sanger sequencing. Pathogenicity was predicted using in silico tools and ACMG criteria. Structural consequences were modeled with AlphaFold3 and Chimera-X. Functional impact was assessed by Western blot and immunofluorescence in transfected 293T cells. RESULTS: All patients presented with neonatal hemolytic anemia. Genetic analysis revealed compound heterozygosity for PKLR mutations. Two novel variants were identified: c.1708G>A (p.Val570Met) and c.1430C>T (p.Thr477Ile). In silico analysis predicted both as damaging. Structural modeling suggested p.Val570Met disrupts inter-subunit hydrogen bonds, while p.Thr477Ile lies within the fructose-1,6-bisphosphate (FBP) binding loop. In vitro, both mutants showed reduced protein expression compared to wild-type but normal cytoplasmic localization. CONCLUSION: We identified two novel PKLR variants expanding the mutational spectrum of PKD. Integrated analysis suggests p.Val570Met may impair tetramer stability, whereas p.Thr477Ile likely affects allosteric regulation. These findings underscore the value of combining clinical phenotyping with functional studies for accurate variant interpretation in PKD.
INTRODUCTION: Myeloproliferative neoplasms (MPNs), encompassing polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF), are hematologic malignancies characterized by recurrent somatic mut...INTRODUCTION: Myeloproliferative neoplasms (MPNs), encompassing polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF), are hematologic malignancies characterized by recurrent somatic mutations. Despite advances in next-generation sequencing (NGS), the genetic basis of thrombotic risk and bone marrow (BM) fibrosis in MPNs remains unclear. METHODS: Patients diagnosed with classical MPN at Trakya University Faculty of Medicine were retrospectively analyzed (2018-2025). Mutation profiling was conducted on BM aspirates using a 78-gene panel on the Illumina NextSeq platform, with clinically relevant variants (VAF ≥ 2%) interpreted via Qiagen Clinical Insight. RESULTS: Among 91 patients with MPN, the most frequent mutations were JAK2 (71.4%), TET2 (23.1%), DNMT3A (15.4%), ASXL1 (11%), and splicing factor mutations (SFMs; 12%). Arterial events occurred in 45.1% of patients and were associated with age (p < 0.001), higher Charlson Comorbidity Index (CCI; p < 0.001), ASXL1 (p:0.019), and SFMs (p:0.009). VTE occurred in 38.5% and was associated with JAK2 status and allele burden (p:0.004 and p:0.012 respectively), TET2 (p < 0.001), and SFMs (p:0.002); the JAK2/TET2 co-mutant subgroup had the highest risk of VTE risk in multivariable analysis (OR: 2.9, 95% CI: 1.4-5.7; p:0.002). Advanced BM fibrosis was associated with SFMs (p:0.018). Increased mutational burden correlated with both venous and arterial thrombosis (p:0.021 and p:0.044, respectively). In survival analyses, SFMs (HR: 5.2; p:0.008), ASXL1 (HR: 3.8; p:0.030), and PMF diagnosis (HR: 3.8; p:0.045) were associated with inferior overall survival (OS). CONCLUSION: Molecular profiling may provide clinically relevant thrombotic risk stratification in classical MPNs. JAK2/TET2 co-mutation was linked to VTE, while ASXL1 and SFMs were associated with adverse phenotypes including vascular events, BM fibrosis, and inferior OS.
Healthcare systems in wealthy countries are facing rising costs due to ageing populations and the prevalence of chronic illnesses, leading to constrained budgets. Pathology services, essential for accurate diagnosis and...Healthcare systems in wealthy countries are facing rising costs due to ageing populations and the prevalence of chronic illnesses, leading to constrained budgets. Pathology services, essential for accurate diagnosis and treatment, have merged to cope with these financial pressures. However, maintaining laboratory quality standards during economic downturns remains critical, as it directly affects patient outcomes, safety and satisfaction. This review explores evidence-based approaches to maintaining quality standards amid financial difficulties. Major challenges include hiring freezes, heavier workloads and an increased risk of burnout, which is associated with higher error rates and staff turnover. The Conservation of Resources theory helps explain burnout by highlighting the importance of organisational support. Strategies to mitigate burnout include policies for digital disconnection, the promotion of transformational leadership among junior managers and the fostering of supportive leadership at higher levels. Additional measures involve sustaining competencies through Entrustable Professional Activities, structured feedback, cross-training and Lean task prioritisation. Initiatives focused on staff well-being, centred on autonomy, psychological empowerment and happiness, are crucial for preserving morale and performance. Overall, these strategies demonstrate that quality can be maintained through structured, evidence-based interventions even during challenging times, thereby ensuring patient-focused care and organisational resilience.
INTRODUCTION: The bone marrow microenvironment plays crucial roles in the pathogenesis of multiple myeloma. Previous studies have shown that there are differences in the metabolomics of bone marrow suspension and plasma,...INTRODUCTION: The bone marrow microenvironment plays crucial roles in the pathogenesis of multiple myeloma. Previous studies have shown that there are differences in the metabolomics of bone marrow suspension and plasma, but there is rare research on bone marrow biopsy tissue. This study aims to establish a new metabolomics approach to analyze bone marrow biopsy tissues and explore its diagnostic value. METHODS: We collected bone marrow biopsy tissues from 19 newly-diagnosed multiple myeloma(MM), 29 monoclonal gammaglobulinemia of undetermined significance(MGUS) and 30 lymphoma without bone marrow invasion as the controls. Metabolomics analysis was conducted using high-performance liquid chromatography-mass spectrometry. Diagnostic Biomarkers and associated pathway were analyzed. RESULTS: The metabolic profiles differed between the controls and MGUS or MM. For MGUS versus MM comparison, Phenylalanyl-Valine and N-Acetylornithine were significantly down-regulated in the MM while N-Acetylornithine has the best diagnostic value with the sensitivity and specificity of 87.5% and 71.4%. For the controls vs. MM comparison, N-Acetylornithine and Aspartyl-Arginine differed markedly. Compared with the controls group, N-Acetylornithine was significantly down-regulated in the MM group, and the AUC of N-Acetylornithine for MM was 0.933, with sensitivity and specificity of (90.0% and 93.3%). N-Acetylornithine displayed the same trends as the comparison between MGUS and MM. Furthermore, when comparing MGUS and MM, Arginine biosynthesis was identified as significantly disrupted in which N-Acetylornithine was involved. CONCLUSIONS: We establish a novel method for metabolomic profiling of bone marrow biopsy tissue and provide evidence of metabolic alterations, particularly reduced N-acetylornithine levels, in MGUS and MM patients.
INTRODUCTION: Bone marrow iron staining is traditionally used to assess iron stores and identify ring sideroblasts (RS), a key feature of sideroblastic anaemia and historically important in the classification of myelodys...INTRODUCTION: Bone marrow iron staining is traditionally used to assess iron stores and identify ring sideroblasts (RS), a key feature of sideroblastic anaemia and historically important in the classification of myelodysplastic and myelodysplastic/myeloproliferative overlap neoplasms. With the adoption of the WHO 2022 classification and increasing availability of next-generation sequencing (NGS), the diagnostic relevance of routine iron staining warrants re-evaluation. METHODS: We retrospectively analysed 101 consecutive adult diagnostic bone marrow examinations (51 myeloid, 50 lymphoid) performed at a regional Australian hospital between January 2023 and June 2024. Perls' Prussian blue staining was routinely performed. Ferritin was available in 76% of cases, and molecular testing in 35% of cases. Two independent reviewers graded bone marrow iron using the Gale score and enumerated RS. Diagnostic impact was categorised as either no impact, supportive but non-essential, or potential diagnostic miss. Interobserver agreement, correlation with serum ferritin, and diagnostic performance for iron deficiency were evaluated. RESULTS: Iron staining had no diagnostic impact in 94% of cases and was supportive but non-essential in 6%; omission would not have altered the final diagnosis in any case. RS ≥ 15% were identified in seven myeloid cases, including MDS, AML, and therapy-related myeloid neoplasms. SF3B1 mutations were present in three RS-positive cases; RS presence did not independently determine diagnosis. Interobserver agreement for iron grading was good (weighted κ = 0.77, 95% CI 0.64-0.89). Correlation between marrow iron grade and serum ferritin was weak (r ≈ 0.38). Low marrow iron grade (≤ 1) showed variable sensitivity (33% to 67%) but high specificity (> 90%) for iron deficiency. CONCLUSION: Routine bone marrow iron staining rarely influences diagnostic classification in contemporary practice. RS morphology is supportive but not determinative, and marrow iron grading correlates poorly with systemic iron status. These data support a targeted rather than routine approach to marrow iron staining, particularly where NGS is available and routinely performed in de novo diagnoses.
INTRODUCTION: Investigator-led research and quality improvement initiatives have led to important improvements in the diagnostic assessment of platelet function disorders (PFD). METHODS: Personal reflections were used to...INTRODUCTION: Investigator-led research and quality improvement initiatives have led to important improvements in the diagnostic assessment of platelet function disorders (PFD). METHODS: Personal reflections were used to summarize our contributions to knowledge on PFD diagnostic assessment, pathogenesis, and bleeding risks. RESULTS: Light transmittance platelet aggregometry (LTA) and whole mount electron microscopy assessment for platelet dense granule deficiency (DGD) both detect abnormalities that are highly predictive of a bleeding disorder. Observations on LTA findings that are predictive of a bleeding disorder (including those specific to certain conditions) have been incorporated into guidelines to reduce LTA interpretation errors. Efforts to improve LTA assessment of PFD with thrombocytopenia (e.g., Bernard Soulier syndrome) have led to validated, trustworthy diagnostic procedures. Commonly encountered PFD that manifest with abnormal aggregation responses to multiple agonists and/or DGD are now established to have significantly increased bleeding risks, emphasizing the need for diagnosis and treatment. Unraveling of the molecular pathogenesis of some specific PFD has provided important insights and simplified diagnosis. In the case of Quebec platelet disorder (QPD), the pathogenesis is a unique gain-of-function defect in fibrinolysis from a mutation that repositions a megakaryocyte-specific enhancer that normally upregulates VCL expression during megakaryopoiesis and "rewires" PLAU, increasing its expression > 100-fold in megakaryocytes only. Presently, the molecular causes of many "commonly encountered" PFD await elucidation. CONCLUSIONS: Research has meaningfully improved diagnostic laboratory testing for PFD. Unraveling the causes of "commonly encountered" PFD will be important to understanding their pathogenesis and increasing the yield of diagnosis by genetic investigations.
INTRODUCTION: The Atellica HEMA 580 is a novel haematology analyser providing eight-part leukocyte differential and optical platelet (PLT) counting. We compared its analytical and clinical performance with that of the AD...INTRODUCTION: The Atellica HEMA 580 is a novel haematology analyser providing eight-part leukocyte differential and optical platelet (PLT) counting. We compared its analytical and clinical performance with that of the ADVIA 2120i. METHODS: A total of 796 residual whole blood specimens were analysed. Carryover, linearity and precision were assessed. Inter-instrument comparison was performed for complete blood counts and five-part differential (5-Diff) parameters using Passing-Bablok regression and Bland-Altman analyses. Additional regression against manual microscopy was conducted for flagged samples, including PLT, reticulocytes and common 5-Diff parameters. RESULTS: Carryover was < 1% for all parameters. Linearity was excellent (R ≥ 0.99). Precision was within acceptable limits; coefficient of variant was < 5% for most parameters, except optical PLT, lymphocytes, monocytes, eosinophils and nucleated red blood cells under specific conditions. Inter-instrument comparisons revealed remarkably high correlations (r ≥ 0.9) for most parameters, whereas mean corpuscular haemoglobin concentration (MCHC) (r = 0.429) and basophils (r = 0.627) showed weaker concordance. Passing-Bablok regression indicated minimal systematic or proportional bias for most parameters. Bland-Altman analysis showed that > 95% of results were within the 95% limits of agreement, except white blood cells, mean corpuscular volume and PLT (94.7%-94.8%). Comparison with microscopy confirmed good concordance for PLT and reticulocytes; however, lymphocytes and monocytes had proportional and systematic biases. CONCLUSION: The Atellica HEMA 580 demonstrated reliable analytical performance and strong agreement with the ADVIA 2120i. This supports its suitability as a routine haematology analyser, providing accurate and clinically comparable results.