BACKGROUND:To develop a machine learning model for early differentiation of primary immune thrombocytopenia (pITP) from connective tissue disease-related thrombocytopenia (CTD-TP) in children presenting with thrombocytopenia. METHOD:A retrospective study was conducted on 387 newly diagnosed children with thrombocytopenia. All patients were clinically diagnosed and divided into a training set and a test set in a 7:3 ratio. Six machine learning algorithms, including XGboost, RF, SVM, LR, GBDT, BPNN, were used to establish differential diagnostic models for pITP and CTD-TP. The model performance was evaluated by accuracy, precision, recall, F1 score, and area under the receiver operating characteristic curve (AUC). Explain the importance and contribution of model feature variables through the Shapley Additive Explanations (SHAP) method. RESULT:A total of 387 patients were included in the study, including 347 cases of pITP and 40 cases of CTD-TP. Based on the above evaluation indicators, the BPNN model had the best performance, with an F1 score of 0.95, an accuracy rate of 94.87%, and an AUC of 0.9738. According to the SHAP value, the top 10 influencing factors were selected as age, erythrocyte sedimentation rate, antinuclear antibodies, IgM, Complement C3, T + B + NK%, B cells, body weight, complement C4, lymphocyte count. CONCLUSION:The successful construction of the pITP and CTD-TP identification models shows that BPNN has the best performance and can provide clinical strategies for early disease identification based on the importance of its variables.
BACKGROUND:Human papillomavirus (HPV)-associated cancers are preventable with vaccination. Serologic assays monitor antibody responses, but their complexity and cost pose limitations when evaluating population level immunity. We developed an L1-based assay for the detection of multiple type-specific HPV antibodies. METHODS:The Luminex-based assay used recombinant L1 proteins from seven HPVs (HPV-6/11/16/18/35/45/58) for the detection of type-specific immunoglobulin (Ig)G and IgA. Cutoffs were established using plasma from infants and children. The assay was optimized and qualified using a sample set from a blinded proficiency panel developed at the Frederick National Laboratory for Cancer Research. RESULTS:Sensitivity and specificity for anti-HPV-6/11/16/18/58 IgG were 100% at the Mean + 3 standard deviation [SD] and Mean + 5 SD thresholds. Sensitivity for HPV-45 was 100% and 92.9% at the +3 SD and +5 SD thresholds, respectively. IgA detection showed moderate positivity among vaccinated individuals, with peak estimates of 96.4% (HPV-11) and 85.7% (HPV-58) at the +3 SD threshold. IgA was correlated with IgG (Spearman's ρ = 0.74). The L1-assay differentiated high, intermediate, low, and negative response groups from the blinded samples for all HPV genotypes (p < 0.0001). CONCLUSION:The L1-binding assay has the potential to be a scalable option for high-throughput detection of broad-spectrum HPV immunoglobulins, but further standardization and validation are needed.
BACKGROUND:Pseudohypercalcemia is easily misdiagnosed in clinical practice, potentially leading to unnecessary interventions. Multiple myeloma (MM) presenting with concurrent hypergammaglobulinemia represents a rare etiology of pseudohypercalcemia, and there are few reports of systematic laboratory validation of this pathological entity. METHODS:We report a patient with MM who presented with markedly elevated serum total calcium on routine biochemical testing. The hypercalcemia persisted despite 1 month of calcitonin therapy administered at an external hospital. After admission, the clinical laboratory team performed a panel of confirmatory assays, including direct ionized calcium measurement, post-dilution linearity evaluation, inductively coupled plasma mass spectrometry (ICP-MS) verification, and polyethylene glycol (PEG) precipitation testing. RESULTS AND CONCLUSIONS:The results confirmed that the elevation in serum total calcium was attributed to enhanced calcium binding to circulating globulins, secondary to hypergammaglobulinemia, rather than analytical interference in the detection system. During clinical follow-up, serum total calcium levels normalized progressively in parallel with the decline in globulin levels following targeted anti-myeloma therapy. This case demonstrates that clinicians and laboratory professionals should maintain a high index of suspicion for pseudohypercalcemia when encountering patients with marked paraproteinemia or abnormal circulating macromolecules. A multimodal differential diagnostic approach using complementary laboratory indicators is warranted to avoid unnecessary examinations and interventions, thereby reducing the waste of medical resources and preventing potential adverse events associated with inappropriate treatment.
BACKGROUND:Hearing loss is one of the most common sensory disorders caused by genetic and environmental factors. Autosomal recessive non-syndromic hearing loss (ARNSHL) is extremely heterogeneous, with over 100 genes known to be involved. Hearing loss can result from mutations in the LOXHD1 gene, which codes a highly conserved protein known as lipoxygenase and is located at 18q21.1. Despite the association between LOXHD1 mutations and ARNSHL, there are still few documented cases. METHODS:We examined a case of non-syndromic hearing loss in an Iranian family with a history of consanguinity and several affected siblings. Next-generation sequencing (NGS) was conducted on the proband to discover causal genetic alterations. Sanger sequencing was employed to confirm the identified variation. RESULTS:A novel likely pathogenic variant in the LOXHD1 gene, c.3713dupA (p.Asp1238Glufs*10), was identified. Sanger sequencing was used to confirm that the affected family members had this frameshift mutation. CONCLUSION:Our findings broaden the mutational range of LOXHD1 linked to ARNSHL. This unique variant enhances the comprehension of the genetic underpinnings of hearing loss and may aid in molecular diagnostics and genetic counseling for impacted families.
BACKGROUND:Deletions involving the 1q43q44 region are among the most recurrent terminal chromosomal imbalances and are associated with a recognizable neurodevelopmental phenotype. These deletions may occur as isolated events or in the context of more complex rearrangements. CASE PRESENTATION:We describe presumed monozygotic twin sisters (patients 1 and 2) with a de novo 1q43q44 deletion and an unrelated patient (patient 3) carrying a similar deletion combined with a 21q22.3 duplication derived from a t(1;21)(q43;q22.3)mat. All three patients exhibited features consistent with the distal 1q43q44 deletion syndrome, including microcephaly, corpus callosum abnormalities, developmental delay, intellectual disability, language impairment, hypotonia, micro-/retrognathia, and congenital heart defects. RESULTS:Cytogenetics and FISH analyses confirmed distal 1q deletions in all patients. Chromosomal microarray analysis identified a ~9.5 Mb deletion in patient 1 (her twin was not tested) and a ~12.1 Mb deletion in patient 3. Despite differences in deletion size and genomic context, both rearrangements disrupted a largely overlapping interval containing 171 genes, including the dosage-sensitive neurodevelopmental genes AKT3, HNRNPU, and ZBTB18 as well as other monoallelic expressed genes (GREM2 and NLRP3) and, in patient 3 only, RYR2. Functional enrichment analyses highlighted pathways related to neurodevelopment, epilepsy, and cardiac function. Although the twins showed similar congenital manifestations, patient 1 later developed seizures whose origin could not be assessed. CONCLUSION:These findings further support the role of haploinsufficient genes within the distal 1q43q44 region, particularly AKT3, HNRNPU, and ZBTB18, to the core neurodevelopmental phenotype and highlight the value of high-resolution genomic analysis for patient characterization.
BACKGROUND:Systemic lupus erythematosus (SLE) is a multifactorial autoimmune disease characterized by multi-organ damage, with genetic, environmental, and viral factors implicated in its pathogenesis. Herpesviruses such as Epstein-Barr virus (EBV) and cytomegalovirus (CMV) are known to trigger or exacerbate SLE, potentially through mechanisms like molecular mimicry and immune dysregulation. CASE PRESENTATION:We report the case of a 10-year-old girl who presented with a malar rash following sun exposure, accompanied by fever and arthritis. Laboratory investigations revealed the presence of multiple autoantibodies and serological evidence of active co-infection with both EBV and CMV. A diagnosis of SLE with mild activity was established. The patient received intravenous methylprednisolone, belimumab, and oral ganciclovir. This led to significant clinical improvement, including resolution of the facial rash and normalization of joint mobility. Follow-up assessments showed near-complete normalization of inflammatory markers and autoantibody titers. However, causality cannot be established from a single case, and proposed mechanisms remain hypotheses. CONCLUSION:This case suggests that acute co-infection with EBV and CMV may play a role in the pathogenesis of SLE, possibly via molecular mimicry and B-cell activation. Therefore, routine screening for EBV and CMV in adolescent SLE patients may be warranted to advance etiological research and inform personalized treatment strategies.
BACKGROUND:Immune-mediated platelet transfusion refractoriness (IPTR) remains a major clinical challenge, with immune alloimmunization against HLA and HPA as the predominant causes. Although HLA cross-reactive groups (CREGs) provide a practical alternative to allele-level matching, methods for simultaneous HLA-CREG and HPA genotyping remain limited despite their potential to improve platelet matching. METHODS:Specific U-anchoring primers (UAPs) and amplification refractory mutation system (ARMS) primers were designed and optimized based on one or more SNPs within HLA-CREG subgroups and HPA alleles. Reaction conditions were optimized, and analytical performance was evaluated. A total of 200 donor DNA samples were tested and compared with NGS and a commercial HPA sequencing kit, with discrepant results verified by Sanger sequencing. RESULTS:We developed and optimized a novel multiplex real-time quantitative PCR (qPCR) assay integrating UAPs and ARMS primers for simultaneous detection of HLA-A/B CREGs and major HPA alleles in 12 reaction tubes. The assay demonstrated an overall limit of detection (LOD) of 0.838 ng/μL and the intra-/inter-assay coefficients of variation were both below 5%, indicating high reproducibility. HLA-CREG typing results were 100% concordant with next-generation sequencing. Sixteen discrepant HPA genotyping results were identified compared with the commercial kit, all of which were confirmed by Sanger sequencing and were consistent with the multiplex qPCR assay. CONCLUSIONS:This multiplex qPCR assay enables rapid, accurate, and cost-effective simultaneous detection of HLA-CREGs and HPA alleles, providing a practical tool for comprehensive platelet compatibility assessment and improved transfusion matching.
BACKGROUND:Sepsis remains difficult to diagnose since blood culture has low sensitivity and current biomarkers lack sufficient specificity. We sought to identify robust diagnostic and therapeutic targets for neonatal sepsis through an integrated bioinformatics and experimental workflow. METHODS:The GSE69686 dataset (108 neonates; 48 sepsis, 60 controls collected ≤ 3 days after birth) was analyzed using a multi-step pipeline: WGCNA to identify sepsis-associated modules; limma-based differential expression; feature selection by LASSO and SVM-RFE; immune deconvolution (CIBERSORT); and pathway interrogation (GSEA). Drug enrichment (Enrichr/DSigDB) and molecular docking (AutoDock) supported targetability. Experimental validation employed a THP-1 sepsis-like model (1 μg/mL LPS, 24 h) with qRT-PCR, shRNA-mediated BST1 knockdown, Western blot, ELISA (IL-6, TNF-α, IL-1β), CCK-8, and Annexin V/PI flow cytometry. RESULTS:Intersecting the outputs of LASSO and SVM-RFE yielded three robust biomarkers-FCER1G, IL1R2, and BST1. These genes are associated with broad shifts in inferred immune infiltration, and GSEA linked their high expression to INFLAMMATORY RESPONSE; BST1 and FCER1G were additionally enriched for TNFα/NF-κB signaling. In THP-1 cells, LPS upregulated all three genes; focusing on BST1, shRNA knockdown partially rescued proliferation (0-72 h), reduced IL-6/TNF-α/IL-1β secretion, and lowered apoptosis relative to sh-NC. Docking analysis suggested artesunate as a candidate modulator with a binding energy of -6.27 kcal/mol to BST1. CONCLUSION:Our integrated pipeline nominates FCER1G, IL1R2, and BST1 as biologically coherent biomarkers of sepsis and highlights BST1 as a plausible therapeutic target warranting further investigation. These findings support external validation in multicenter cohorts and mechanistic extension-particularly functional testing of predicted modulators-in primary cells.
BACKGROUND:In a context of HIV seropositivity below 5%, transitioning from a two- to a three-test algorithm is required to secure positive predictive values > 99% for mitigating false reactivity. With Cameroon having 2.7% seroprevalence, we sought to ascertain events of false reactivity from eight WHO-prequalified HIV rapid diagnostic tests (RDTs) and design suitable combinations for the country three-test algorithm. METHODS:A facility-based study was conducted in July-August 2021 among 172 consenting clients attending health facilities in Yaoundé-Cameroon. Whole blood was collected and samples characterized in parallel using KHB Diagnostic Kit for HIV (1 + 2) Antibody and INNO-LIA HIV I/II Ag/Ab to rule out reactive cases. Two lots of each selected RDT were tested simultaneously in quadruplicate on HIV non-reactive samples and repeated by a different tester. RDTs were considered false positive if any reactivity was detected. RESULTS:Following characterization, 25 samples were HIV-reactive (i.e., ruled out) and 147 non-reactive. Following random selection, 100 non-reactive samples tested for false reactivity revealed: 0% (0/100) First Response HIV 1-2-O Card, 0% (0/100) First Response HIV1 + 2/Syphilis Combo, 0% (0/100) STANDARD Q HIV/Syphilis Combo, 0% (0/99) ONE STEP Anti-HIV1&2 (one invalid result); 1% (1/100) Determine HIV Early Detect and Bioline HIV/Syphilis Duo, 2% (2/100) Alere Determine HIV-1/2, 4% (4/100) Rapid Test for Antibody to HIV. CONCLUSION:In the Cameroonian context, HIV false reactivity ranged from 0% to 4% across selected RDTs. Thus, a reliable three-test algorithm ranks First Response HIV 1-2-0 Card, First Response HIV1 + 2/Syphilis Combo or STANDARD Q as suitable RDT-1 to mitigate false-positive events at the country-level.
OBJECTIVE:The aim of this study was to investigate the relationship between human epididymis protein 4 (HE4) levels in seminal plasma and semen liquefaction in males. METHODS:A total of 314 healthy men from the andrology department were included. The correlations between HE4 levels and prostate-specific antigen (PSA) levels as well as conventional semen parameters (sperm density, the proportion of non-progressively motile spermatozoa, seminal liquefaction time) were analyzed, and logistic regression was further employed to examine the association between HE4 levels and the risk of poor semen liquefaction. RESULTS:The level of HE4 in seminal plasma was negatively correlated with semen liquefaction time (r = -0.207, p < 0.001) and positively correlated with seminal plasma PSA levels (r = 0.280, p < 0.001), suggesting a potential link to the process of semen liquefaction. Continuous HE4 levels were standardized by dividing by 100, and participants were then categorized into quartiles (Q1-Q4) based on the standardized HE4 values. After adjusting for confounding factors, the risk of poor semen liquefaction in the Q3, Q2, and Q1 groups of the lower HE4 level was 4.024 times (95% CI: 1.908-8.485; p < 0.001), 6.320 times (95% CI: 2.818-14.175; p < 0.001), and 5.190 times (95% CI: 2.217-12.15; p < 0.001) higher, respectively, compared to the highest HE4 quartile (Q4). CONCLUSION:This suggests that a low level of HE4 in seminal plasma is an independent risk factor for poor semen liquefaction.
BACKGROUND:Chronic kidney disease progression is influenced by anemia and comorbidities; however, the possibility of a real effect of these two phenomena has hardly been the subject of research. OBJECTIVE:The present study set out to evaluate the association between anemia and burden of comorbidities on CKD progression and hospitalizations over 1 year. METHODS:A retrospective study involving 120 adult CKD patients. Anemia was defined as hemoglobin < 13 g/dL in males and < 12 g/dL in females while comorbidity burden was defined by a simple comorbidity index: hypertension, diabetes, and cardiovascular disease. CKD progression was coded as a ≥ 25% decline in eGFR or stage conversion. Multivariate logistic regression identified predictors of hospitalization. RESULTS:Participants' mean age was 58.3 years, and 53.3% were males. Anemia was present in 67.5% of the cases, and 33.3% had two or more comorbidities. CKD progression was seen in 43.3% at 1 year; patients with anemia and two or more comorbidities showed the highest progression rate at 77.5% (p < 0.001). Progressors had significantly lower hemoglobin and higher RDW values. Hospitalization was documented in 29.2%, and anemia independently predicted hospitalization (OR 3.4); comorbidity index ≥ 2 (OR 2.8), and RDW > 14.5% (OR 2.6). Each 1 g/dL increase in hemoglobin reduced hospitalization risk by 28% (OR 0.72). CONCLUSION:Anemia and multiple comorbidities jointly increased the risk of CKD progression and hospitalization. Routine hematological parameters along with assessing comorbidities might have a role to play in early risk stratification and guiding focused interventions in CKD management.
BACKGROUND:Autoverification improves laboratory efficiency by reducing manual reviews; however, rule-based systems may lack flexibility, particularly in complete blood count (CBC) testing. Machine learning (ML) can potentially enhance performance by recognizing complex patterns in laboratory data. Therefore, we aimed to evaluate ML-based autoverification systems in classifying CBC results compared to traditional rule-based methods. METHODS:This cross-sectional study was conducted at the Hematology Unit of Songklanagarind Hospital, a tertiary care center in southern Thailand. In total, 63,201 CBC results from the Sysmex XN-10 analyzer (January-December 2024) were analyzed. The samples were categorized as verifiable or non-verifiable according to predefined criteria. The data were split 80:20 into training and testing sets. The Random Forest, Decision Tree, and Extreme Gradient Boost models were trained and evaluated against the rule-based system of the hospital using sensitivity, specificity, positive predictive value, and negative predictive value. The feature importance was analyzed for the best-performing model. RESULTS:Of all the results, 43,661 (69.1%) were verifiable and 19,540 (30.9%) were non-verifiable, mainly because of white blood cell abnormalities, platelet clumps, or differential discrepancies. In the 12,641-sample test set, all the ML models outperformed the rule-based system. XGBoost achieved the best balance, with a sensitivity of 95%-98% and a specificity of 68%-81% at recall thresholds of 95% and 97.5%, respectively. CONCLUSION:ML demonstrated improved performance compared with rule-based autoverification. Although ML may reduce unnecessary manual reviews, validation is required in different settings before clinical adoption.
BACKGROUND:Minimal residual disease (MRD) assessment by next-generation flow cytometry (NGF) is an important prognostic tool in multiple myeloma (MM). Although the EuroFlow NGF assay provides highly standardized and sensitive MRD detection, its dual-tube workflow requires larger sample volumes and longer processing time, potentially limiting efficiency in routine clinical practice. This prospective study evaluated a 10-color single-tube (ST-10C) MRD assay by directly comparing its analytical and clinical performance with the EuroFlow NGF assay. METHODS:MRD analysis was performed in 40 patients with MM, of whom 39 who achieved very good partial response or complete response were included in the final analysis. Each sample was analyzed in parallel using both assays. Analytical sensitivity, qualitative and quantitative agreement, and progression-free survival (PFS) were assessed. RESULTS:Both assays achieved analytical sensitivity below 10-5. MRD was detected in 48.7% of samples by the ST-10C assay and in 38.5% by the EuroFlow NGF assay, with substantial agreement (κ = 0.79). Concordance remained high using the clinical MRD cutoff (≥ 0.001%) (κ = 0.84). Quantitative MRD levels showed strong correlation between the assays (ρ = 0.964) without significant systematic bias. MRD-positive patients showed a trend toward inferior PFS compared with MRD-negative patients by both assays. CONCLUSIONS:The ST-10C assay demonstrated analytical performance comparable to the EuroFlow NGF assay, supporting its adoption as a more efficient and streamlined alternative for routine MRD assessment in MM.
OBJECTIVE:This study aimed to compare the diagnostic accuracy of four serum biomarkers-carbohydrate antigen 125 (CA125), human epididymis protein 4 (HE4), the Risk of Ovarian Malignancy Algorithm (ROMA), and the Copenhagen Index (CPH-I)-for the preoperative discrimination of benign/borderline from malignant ovarian cancer, with particular attention to menopausal status. METHODS:A retrospective study enrolled 345 patients with histologically confirmed ovarian tumors (187 benign/borderline; 158 malignant). Serum CA125 and HE4 were measured and reported by the electrochemiluminescence immunoassays, along with the calculated indices of ROMA and CPH-I. Diagnostic performance was assessed by the area under the curve (AUC), sensitivity, and specificity, respectively. RESULTS:Among the 158 ovarian tumors patients, CPH-I demonstrated superior discriminative capacity with the highest AUC (0.924), followed by ROMA (0.916), HE4 (0.897), and CA125 (0.865). Menopausal stratification identified CPH-I as optimal for premenopausal women (AUC 0.886) and ROMA for postmenopausal women (AUC 0.928). ROMA exhibited the highest overall sensitivity (84.8%), whereas CA125 achieved the highest overall specificity (94.1%). In premenopausal patients, HE4 demonstrated exceptional specificity (98.6%), while postmenopausal women showed equivalent specificity between HE4 and CPH-I (both 97.8%). In combined panels, CA125 + HE4 + CPH-I yielded the highest overall accuracy (AUC 0.925), and CA125 + HE4 + ROMA provided optimal postmenopausal performance (AUC 0.930). CONCLUSIONS:CPH-I demonstrated favorable diagnostic performance compared with conventional biomarkers. Our findings advocate for a menopausal status-guided clinical algorithm, utilizing a CPH-I-based panel for premenopausal women and a ROMA-based panel for postmenopausal women to maximize diagnostic precision.
BACKGROUND:To evaluate the detection capabilities of prenatal screening and diagnostic methods for fetal chromosomal mosaicism and analyze associated pregnancy outcomes. METHODS:This retrospective study included 79 cases of fetal chromosomal mosaicism identified by karyotyping among 8106 women undergoing prenatal diagnosis. Sensitivity of screening methods (first-trimester serum screening, second-trimester serum screening, and noninvasive prenatal testing (NIPT)) and concordance of diagnostic methods (chromosomal microarray analysis (CMA), multiple STR locus analysis technique (QF-PCR)) with karyotyping were assessed. Pregnancy outcomes were analyzed. RESULTS:NIPT demonstrated significantly higher sensitivity (90.74%) compared with first-trimester serum screening (57.41%) and second-trimester serum screening (39.29%) (p < 0.001). CMA showed high concordance with karyotyping (90.00%), whereas QF-PCR exhibited significantly lower concordance (58.21%) (p < 0.001). The pregnancy termination rate was 57.14% (40/70). All trisomy 21 and trisomy 13 mosaicism cases resulted in termination, while decisions regarding other autosomal and sex chromosome mosaicism were more variable. CONCLUSION:NIPT outperforms traditional serum screening for detecting chromosomal mosaicism, though karyotyping remains essential for confirmation. CMA aligns well with karyotyping, particularly for low-level mosaicism. The high termination rate underscores the critical need for multidisciplinary genetic counseling integrating genetic, imaging, and clinical data.
BACKGROUND:Acute kidney injury (AKI) affects approximately 30%-60% of intensive care unit (ICU) admissions, but early detection remains difficult because creatinine- and urine-output-based KDIGO criteria are delayed and confounded in critical illness. OBJECTIVE:To summarize the biomarker landscape for ICU-associated AKI, focusing on kidney-targeted injury/stress proteins and circulating cell-free DNA (cfDNA), including mitochondrial DNA (mtDNA). METHODS:PubMed/MEDLINE and Embase were searched from inception to December 31, 2025, using terms related to AKI, critical illness, NGAL, KIM-1, IL-18, L-FABP, [TIMP-2]·[IGFBP7], cfDNA, mtDNA, and damage-associated molecular patterns. Adult ICU studies reporting diagnostic performance, risk stratification, or clinically relevant outcomes were prioritized, especially in sepsis, cardiac surgery, shock, and trauma. RESULTS:NGAL, KIM-1, IL-18, L-FABP, and [TIMP-2]·[IGFBP7] often rise before creatinine or urine-output changes, providing early kidney-proximal signals of tubular injury or stress. cfDNA/mtDNA instead reflects systemic cell death, mitochondrial damage, and innate immune activation, capturing multi-organ injury and adding prognostic information beyond clinical scores and creatinine. Across ICU phenotypes, combining clinical risk, an early kidney-targeted marker, and cfDNA/mtDNA may improve identification of patients at risk for severe AKI, renal replacement therapy, and death. CONCLUSIONS:Multimarker strategies may widen the diagnostic window and sharpen prognostication in ICU-associated AKI. Routine implementation will require standardized pre-analytical handling, assay harmonization, biomarker-guided pragmatic trials, and trajectory-based decision tools that demonstrate clinical benefit.
BACKGROUND:Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal malignancy that is often diagnosed after curative treatment is no longer feasible. Existing biomarkers, particularly CA19-9, have limited sensitivity and specificity. Plasma proteins that capture tumor-associated biological alterations may therefore provide useful signals for earlier detection. METHODS:Plasma samples from 99 patients with PDAC and 30 healthy controls were analyzed using data-independent acquisition (DIA) proteomics. Differentially expressed proteins were identified using predefined statistical thresholds and further examined by functional enrichment analysis. Selected candidate biomarkers were validated by ELISA in an independent subset. RESULTS:Among 565 quantified plasma proteins, 52 were differentially expressed between PDAC and controls. These proteins were enriched in extracellular processes, cholesterol metabolism, complement and coagulation cascades, and pancreatic secretion pathways. ELISA validation confirmed higher plasma levels of Cathepsin S, CTRB2, MARCO, PIGR, PRDX6, REG1A, Trypsin-2, and PEP-FAP in patients with PDAC compared with healthy controls. ROC analyses showed moderate-to-good discriminatory performance for several candidates, and the MARCO + PEP-FAP model improved classification compared with either marker alone. CONCLUSION:These findings reveal circulating proteins linked to key PDAC-related biological processes and identify eight candidates for further evaluation in multi-protein diagnostic panels. Larger validation studies incorporating clinically relevant disease control groups are warranted to determine their diagnostic specificity and clinical utility.
BACKGROUND:Measurement of faecal haemoglobin (f-Hb) using the faecal immunochemical test (FIT) in population screening for colorectal cancer (CRC) has reduced cancer-related incidence and mortality. However, temperature-induced f-Hb degradation can decrease FIT accuracy. This study assessed the f-Hb preservation capacity of a new FIT Hb-stabilising buffer at different temperatures and durations. METHODS:Fresh whole faecal samples from individuals with active colorectal bleeding were used for collection of samples using the OC-Sensor FIT (EIKEN CHEMICAL CO. LTD, Japan) containing either the standard (SOC3) or new formulation (SOC4) buffer. Pooled faeces-buffer mixtures (n = 29) were divided into aliquots for incubation at 4°C, 23°C, 35°C, 45°C, and 50°C and analysed for f-Hb concentration at days 0, 3, 7, 10, 14, 21, and 28. Relative f-Hb (percentage change of day 0) in both buffer types was compared between temperatures at each timepoint. Generalised linear modelling identified variables influencing f-Hb degradation. RESULTS:SOC3 exhibited significant f-Hb decline from day 10 at 23°C and from day 3 at all higher temperatures assessed (p < 0.05). In contrast, f-Hb concentrations remained stable within SOC4 for up to 28 days at 23°C, with reductions observed after 7, 7, and 14 days at 50°C, 45°C, and 35°C, respectively (p < 0.05). Higher temperatures and longer storage duration significantly decreased f-Hb concentration, while SOC4 demonstrated superior f-Hb preservation relative to SOC3 (generalised linear model estimate = 15.4, p < 0.001). CONCLUSION:The SOC4 FIT buffer improves f-Hb stability compared with the current formulation and may improve CRC screening accuracy, particularly in high-temperature environments.
PURPOSE:To assess the feasibility of using the cell-free DNA barcode-enabled single-molecule test (cfBEST) for non-invasive prenatal testing (NIPT) of α-thalassemia and β-thalassemia. METHODS:Seventy two thalassemia carrier families participated in the study. The cfBEST method was employed to identify thalassemia genotypes in fetal cell-free DNA extracted from maternal plasma, targeting four α-thalassemia gene mutations and 13 common β-thalassemia gene mutations. Validation was conducted using gap-PCR and PCR-RDB through invasive prenatal diagnosis. All prenatal diagnosis results were later confirmed for accuracy. RESULTS:cfBEST successfully identified 94.6% (88/93) of fetal alleles, achieving a sensitivity of 94% (95% CI, 83.45%-98.75%) and a specificity of 95.35% (95% CI, 84.19%-99.43%). All prenatal diagnosis results were followed up, and the follow-up findings were consistent with the prenatal diagnosis results. CONCLUSION:The findings indicate that cfBEST is a reliable, precise, simple, and cost-efficient method suitable for non-invasive prenatal testing of α-thalassemia and β-thalassemia.
