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15 pages, 1885 KB

Open AccessArticle

One Target, Different Results: The Clinical Impact of Diagnostic Kit Choice in BCR::ABL1 Testing for Chronic Myeloid Leukemia

by Mirjana Suver Stević, Vlatka Periša, Karla Vujičić, Saška Marczi, Jasminka Sinčić-Petričević and Danijela Mjeda

Diagnostics 2026, 16(14), 2216; https://doi.org/10.3390/diagnostics16142216 - 15 Jul 2026

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Abstract

Background: Quantitative PCR measurement of BCR::ABL1 is essential for monitoring molecular response and detecting relapse in chronic myeloid leukemia (CML) patients. Given the availability of multiple commercial kits for cDNA synthesis and minimal residual disease assessment, analytical accuracy and reliability are [...] Read more.

Background: Quantitative PCR measurement of BCR::ABL1 is essential for monitoring molecular response and detecting relapse in chronic myeloid leukemia (CML) patients. Given the availability of multiple commercial kits for cDNA synthesis and minimal residual disease assessment, analytical accuracy and reliability are critical. This study evaluated two commercial RT and qPCR kits for BCR::ABL1 quantification and fusion transcript variant identification. Methods: Total RNA was isolated from peripheral blood, bone marrow, and external quality control samples from the UK NEQAS for Leucocyte Immunophenotyping program. cDNA synthesis was performed using two kits: AffinityScript (ASK) and RT Kit (RTK). BCR::ABL1 transcript levels were determined using the ipsogen^® BCR-ABL1 Mbcr IS-MMR Kit (IPS) and the LightMix^® bcr-abl t(9;22) M/m/µ Kit (TMB). Results were compared with UK NEQAS LI reference data. Fusion transcript variants were analyzed using nested PCR and a commercial qPCR assay. Results: Substantial variability was observed between the TMB and IPS assays, with moderate, non-significant correlation and wide limits of agreement. Established discrepancies resulted in different classifications of molecular response, and IPS results showed better concordance with external quality assessment data. ABL1 quantification revealed significantly higher copy numbers with the RTK compared to the ASK (p < 0.0001), enabling more reliable assessment of deep molecular responses. Statistical analyses indicated systematic and proportional bias between the methods. For variant detection, nested PCR demonstrated higher specificity, while the commercial assay showed limited discriminatory capability. Conclusions: Significant methodological differences may affect clinical interpretation, underscoring the importance of validation and standardization in CML molecular monitoring. Full article

(This article belongs to the Special Issue Advances in Laboratory Analysis and Diagnostics)

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10 pages, 971 KB

Open AccessArticle

Selective Inhibition of Insulin-Degrading Enzyme Eliminates Hemolysis Interference in Serum Insulin Measurements

by María Rodríguez-García, Bernardino González de la Presa, Aleix B. Fabregat-Bolufer, Naira Rico, Helena Castella, Alejandro Calvera-Rayo, Marga Giménez, Felicia A. Hanzu, Manuel Morales-Ruiz and Gregori Casals

Diagnostics 2026, 16(12), 1927; https://doi.org/10.3390/diagnostics16121927 - 22 Jun 2026

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Abstract

Objectives. Hemolysis significantly interferes with insulin measurements in clinical settings, leading to inaccurate results. Although the activity of insulin-degrading enzyme (IDE) is assumed to be the primary mechanism, the potential involvement of additional mechanisms remains unclear. This study aims to determine if [...] Read more.

Objectives. Hemolysis significantly interferes with insulin measurements in clinical settings, leading to inaccurate results. Although the activity of insulin-degrading enzyme (IDE) is assumed to be the primary mechanism, the potential involvement of additional mechanisms remains unclear. This study aims to determine if IDE is the sole cause of this interference by using a selective IDE inhibitor, 6bK, and to explore whether this inhibition can completely prevent hemolysis-related inaccuracies in insulin assays. Methods. The effects of 6bK on insulin degradation were evaluated in hemolyzed and non-hemolyzed serum samples at room temperature, following the CLSI guidelines EP07-A2 and C56-A. Insulin levels were measured using chemiluminescent immunoassays. Additional assessments included the impact of 6bK on serum C-peptide, proinsulin, and standard biochemical parameters. The effects of 6bK were also evaluated at 4 °C and after 21 days of storage at room temperature prior to use. Results. Hemolysis caused a significant decrease in insulin concentrations, dependent on hemolysate levels and incubation time. The addition of 10 µM 6bK completely reversed hemolysis-induced insulin degradation in serum across a broad range of insulin baseline concentrations and degrees of hemolysis. Furthermore, 6bK did not affect insulin levels in non-hemolyzed samples or alter the quantification of C-peptide, proinsulin, or standard biochemical parameters. Conclusions. The decrease in serum insulin concentration due to hemolysis is exclusively attributed to the action of IDE. Selective inhibition of IDE by 6bK effectively eliminates hemolysis-induced interference in insulin measurements, providing a novel and reliable solution for accurate insulin quantification in hemolyzed clinical samples. Full article

(This article belongs to the Special Issue Advances in Laboratory Analysis and Diagnostics)

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17 pages, 1338 KB

Open AccessArticle

Comparative Performance Analysis of Commercial SARS-CoV-2 RNA Detection Assays: Implications for Sensitivity, Specificity, Accuracy, and Diagnostic Response Time

by Adriana Guimarães dos Santos, José Rodrigo Santos Silva, Maria Luísa Rodrigues Nolasco and Marcus Vinicius de Aragão Batista

Diagnostics 2026, 16(10), 1554; https://doi.org/10.3390/diagnostics16101554 - 20 May 2026

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Abstract

Background/Objectives: In 2020, the world found itself in the midst of the SARS-CoV-2 pandemic. The virus has spread globally, resulting in over 779 million cases worldwide. In response to this crisis, there arose a critical need for diagnostic techniques capable of meeting the [...] Read more.

Background/Objectives: In 2020, the world found itself in the midst of the SARS-CoV-2 pandemic. The virus has spread globally, resulting in over 779 million cases worldwide. In response to this crisis, there arose a critical need for diagnostic techniques capable of meeting the overwhelming global demand, including RT-qPCR as the gold standard due to its high sensitivity and specificity. However, RT-qPCR has its limitations, including susceptibility to factors such as inadequate sample collection, variations in viral load, and insufficient clinical validation, all of which can lead to false negatives. Consequently, this study aims to evaluate the clinical performance of four commercial RT-qPCR kits for detecting SARS-CoV-2. Methods: The study utilized 200 nasopharyngeal swab samples collected in January 2022, comparing kits from Qiagen, Seegene, Bio-Manguinhos, and IBMP. Results: Results indicated significant differences in kit performance, with 66% of samples showing consistent results across all kits, and 34% showing discrepancies. Ct values were also analyzed, and statistical tests highlighted varying sensitivities among the kits, ranging from 100% to 86.82%. Conclusions: The study underscores how extraction and purification processes, kit quality, and target gene adequacy critically influence kit performance, influencing the occurrence of false positives and negatives. Full article

(This article belongs to the Special Issue Advances in Laboratory Analysis and Diagnostics)

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15 pages, 2508 KB

Open AccessArticle

Thrombin Generation in Acute and Chronic Liver Disease in Children

by Giovina Di Felice, Anna Lisa Montemari, Andrea Pietrobattista, Luca Della Volpe, Antonella Mosca, Daniela Liccardo, Simona Pezzi, Chiara Giorni, Matteo Luciani, Danilo Alunni Fegatelli, Annarita Vestri and Ottavia Porzio

Diagnostics 2026, 16(9), 1328; https://doi.org/10.3390/diagnostics16091328 - 28 Apr 2026

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Abstract

Background: Pediatric liver disease is frequently associated with abnormal conventional coagulation tests; however, prothrombin time expressed as international normalized ratio (PT-INR) incompletely reflect global hemostatic balance. Thrombin generation assay (TGA) provide an integrated assessment of coagulation and may offer complementary information in children [...] Read more.

Background: Pediatric liver disease is frequently associated with abnormal conventional coagulation tests; however, prothrombin time expressed as international normalized ratio (PT-INR) incompletely reflect global hemostatic balance. Thrombin generation assay (TGA) provide an integrated assessment of coagulation and may offer complementary information in children with acute liver failure (ALF) and chronic liver disease (CLD). Methods: We enrolled 61 pediatric patients with liver disease (50 CLD, 8 ALF, 3 extrahepatic portal vein obstruction EHPVO) and 51 healthy controls. Platelet-poor plasma was prepared according to international recommendations. Thrombin generation was measured using ST Genesia (STG) with normalization to reference plasma. Group comparisons were performed using non-parametric tests; correlations between PT-INR and thrombin generation parameters were assessed, and principal component analysis (PCA) was used to explore the variance structure of thrombin generation indices and conventional coagulation variables. Results: PT-INR was significantly higher in patients than controls, particularly in ALF. Bleeding events were uncommon. Compared with controls, patients showed reduced levels of fibrinogen and multiple procoagulant/anticoagulant factors (including antithrombin and protein C), with increased factor VIII. Among thrombin generation parameters, the endogenous thrombin potential (ETP) ratio differed significantly across groups (p = 0.001), while correlations between PT-INR and thrombin generation parameters were weak or absent, no significant associations were observed even at higher Pediatric/Model for End-Stage Liver Disease scores. PCA separated thrombin generation indices from PT-INR and conventional coagulation factors, suggesting complementary information. Conclusions: In pediatric liver disease, PT-INR does not reliably reflect global coagulation capacity. Thrombin generation testing provides additional, integrative information on hemostasis and may improve laboratory assessment beyond conventional tests. Full article

(This article belongs to the Special Issue Advances in Laboratory Analysis and Diagnostics)

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11 pages, 1506 KB

Open AccessArticle

External Quality Assessment of Molecular Testing for HLA-B*58:01 Allele in Shanghai

by Jing Quan, Pengyin Zhang, Yanqun Xiao, Xiaobo Hu and Yun Bao

Diagnostics 2026, 16(7), 1005; https://doi.org/10.3390/diagnostics16071005 - 27 Mar 2026

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Abstract

Background/Objectives: Allopurinol, a first-line drug for gout and hyperuricemia, carries a risk of severe cutaneous adverse reactions (SCARs). Studies have established a strong association between HLA-B*58:01 and this adverse reaction. Although pre-treatment genotyping is recommended, the reliability of HLA-B*58:01 genetic testing varies across [...] Read more.

Background/Objectives: Allopurinol, a first-line drug for gout and hyperuricemia, carries a risk of severe cutaneous adverse reactions (SCARs). Studies have established a strong association between HLA-B*58:01 and this adverse reaction. Although pre-treatment genotyping is recommended, the reliability of HLA-B*58:01 genetic testing varies across laboratories. This study aims to assess the performance of HLA-B*58:01 genetic testing of clinical laboratories in Shanghai through an External Quality Assessment (EQA) program, evaluating accuracy and standardization. Methods: The EQA program was carried out twice a year in 2023 and 2024. Each EQA sample panel consisted of five distinct samples, including HLA-B*58:01 allele-positive and -negative cell cultures. Sample panels were distributed to clinical laboratories through the cold chain system and results were analyzed and scored. Results: EQA samples used in this study were optimized for evaluating current HLA-B*58:01 genotyping assays, and the EQA samples were proved to be homogeneous and stable through each EQA period. In 2023, 17 and 16 clinical laboratories participated in the two EQA schemes; in 2024, 34 and 33 laboratories participated. A total of 14/17 (82.4%), 16/16 (100%), 33/34 (97.1%), and 33/33 (100%) laboratories achieved “optimal” scores. Conclusions: EQA results indicate that most of clinical laboratories in Shanghai exhibit constantly satisfactory performance for HLA-B*58:01 genotyping. However, a few laboratories still need further improvement. Additionally, EQA has demonstrated to be an important method for monitoring clinical laboratories’ performance. Full article

(This article belongs to the Special Issue Advances in Laboratory Analysis and Diagnostics)

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16 pages, 2294 KB

Open AccessArticle

Second-Trimester Fibrinogen-to-Albumin Ratio and Platelet Activation Markers Compared with the HALP Score for Predicting Preeclampsia

by Cagla Bahar Bulbul and Betul Yakistiran

Diagnostics 2026, 16(4), 613; https://doi.org/10.3390/diagnostics16040613 - 19 Feb 2026

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Abstract

Objective: This study aimed to evaluate the predictive value of second-trimester hemoglobin levels, the hemoglobin–albumin–lymphocyte–platelet (HALP) index, the fibrinogen-to-albumin ratio (FAR), and selected coagulation and platelet activation markers for the development of preeclampsia. Methods: This retrospective cohort study included 262 pregnant women, comprising [...] Read more.

Objective: This study aimed to evaluate the predictive value of second-trimester hemoglobin levels, the hemoglobin–albumin–lymphocyte–platelet (HALP) index, the fibrinogen-to-albumin ratio (FAR), and selected coagulation and platelet activation markers for the development of preeclampsia. Methods: This retrospective cohort study included 262 pregnant women, comprising 131 women who developed preeclampsia and 131 normotensive controls, followed at a tertiary referral center between 2022 and 2023. Maternal demographic, clinical, and laboratory data were obtained from routine second-trimester (14–28 weeks) antenatal assessments. HALP and FAR were calculated using standardized formulas. Group comparisons were performed using appropriate parametric or nonparametric tests. Discriminative performance was assessed using receiver operating characteristic (ROC) curve analysis with bootstrap resampling. Univariate and multivariable logistic regression models were constructed to evaluate independent associations, and combined biomarker models were compared using DeLong’s test. Results: Women with preeclampsia demonstrated significantly lower hemoglobin, hematocrit, platelet count, and albumin levels, alongside higher fibrinogen, D-dimer, LDH, CRP, and platelet activation indices (MPV, PDW, and P-LCR) (all p < 0.05). Both HALP and FAR were significantly higher in the preeclampsia group; however, FAR exhibited superior discriminatory ability (AUC 0.682; 95% CI 0.618–0.751) compared with HALP (AUC 0.619; 95% CI 0.551–0.680). In multivariable analysis, FAR remained a strong independent predictor of preeclampsia (adjusted OR 1.263; 95% CI 1.167–1.368), whereas HALP showed a weaker association. Among combined models, FAR plus platelet distribution width (PDW) provided the highest discrimination (AUC 0.820), significantly outperforming FAR alone (DeLong p = 0.030). Conclusion: While the FAR + PDW model demonstrated improved discriminatory performance, these findings should be interpreted as preliminary. Prospective multicenter studies and external validation are necessary before such biomarkers can be considered for routine clinical use. Full article

(This article belongs to the Special Issue Advances in Laboratory Analysis and Diagnostics)

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13 pages, 4732 KB

Open AccessArticle

Claudin-4 Overexpression Predicts Poor Survival and Platinum Resistance in Epithelial Ovarian Cancer: A Potential Biomarker for Clinical Decision-Making

by Özlem Kutlu, Damla Günenç, Duygu Ayaz, Özlem Özdemir, Halil Taşkaynatan, Celal Akdemir and Muzaffer Sancı

Diagnostics 2025, 15(24), 3163; https://doi.org/10.3390/diagnostics15243163 - 11 Dec 2025

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Abstract

Background/Objectives: Epithelial ovarian cancer (EOC) is a leading cause of death among forms of gynecologic cancer. Significant causes of mortality include high recurrence rates and the development of resistance to platinum-based chemotherapy. This highlights the need for reliable prognostic biomarkers to improve [...] Read more.

Background/Objectives: Epithelial ovarian cancer (EOC) is a leading cause of death among forms of gynecologic cancer. Significant causes of mortality include high recurrence rates and the development of resistance to platinum-based chemotherapy. This highlights the need for reliable prognostic biomarkers to improve patient stratification and inform treatment decisions. Claudin-4, a tight junction protein frequently overexpressed in epithelial tumors, has been associated with tumor progression and resistance to chemotherapy. Methods: We retrospectively analyzed 83 patients with EOC who underwent debulking surgery. Claudin-4 expression was assessed by immunohistochemistry and categorized as high or low based on a semi-quantitative scoring system. Survival outcomes were evaluated using Kaplan–Meier analysis and Cox regression. Predictors of platinum resistance were examined using logistic regression. Results: High Claudin-4 expression was observed in 55.4% of cases and was associated with significantly shorter disease-free survival (DFS) (23 vs. 66 months, p = 0.00024) and overall survival (OS) (85 months vs. NR, p = 0.0031). In multivariable analysis, platinum resistance (DFS; HR 4.99, OS; HR 4.27) and high Claudin-4 expression (DFS; HR 2.46, OS; HR 3.59) were independent predictors of poor outcomes. Logistic regression further demonstrated that high Claudin-4 expression and interval debulking surgery were independent predictors of platinum resistance. Conclusions: High Claudin-4 expression was associated with inferior survival and an increased risk of platinum resistance in EOC. Our findings suggest that Claudin-4 may serve as a negative prognostic biomarker and a potential therapeutic target. Future prospective studies are warranted to further elucidate the underlying mechanisms and validate Claudin-4’s clinical utility. Full article

(This article belongs to the Special Issue Advances in Laboratory Analysis and Diagnostics)

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11 pages, 1003 KB

Open AccessArticle

Reference Intervals and Cut-Off Values for Thyroid Tests in the Croatian Adult Population on the Snibe MAGLUMI X6 Immunoassay Analyzer

by Ivana Lapić, Dragana Šegulja, Željkica Jakoplić, Iva Lukić and Dunja Rogić

Diagnostics 2025, 15(18), 2360; https://doi.org/10.3390/diagnostics15182360 - 17 Sep 2025

Cited by 3 | Viewed by 2253

Abstract

Background/Objectives: To establish reference intervals (RIs) and cut-off values for thyroid-related tests on the MAGLUMI X6 immunoassay analyzer (Snibe Diagnostic, Shenzhen, China) in an adult Croatian population. Methods: This study included 305 healthy individuals who underwent regular preventive medical checkup. The following [...] Read more.

Background/Objectives: To establish reference intervals (RIs) and cut-off values for thyroid-related tests on the MAGLUMI X6 immunoassay analyzer (Snibe Diagnostic, Shenzhen, China) in an adult Croatian population. Methods: This study included 305 healthy individuals who underwent regular preventive medical checkup. The following tests were determined in serum: thyroid-stimulating hormone (TSH), total triiodothyronine (TT3), total thyroxine (TT4), free triiodothyronine (FT3), free thyroxine (FT4), thyroglobulin (Tg), reverse triiodothyronine (revT3), total binding capacity of thyroglobulin (T-uptake), thyroglobulin antibodies (anti-Tg), anti-thyroid peroxidase antibodies (anti-TPO) and thyroid receptor antibodies (TRAb). TSH, TT3, TT4, FT3, FT4, Tg, revT3 and T-uptake results were used for calculating double-sided 95% RIs between the 2.5th and 97.5th percentiles. For anti-Tg, anti-TPO and TRAb, right-sided cut-offs that correspond to the 95th percentile were determined. Results: Reference intervals for TSH, TT4, FT3, FT4, Tg, T-uptake and revT3 did not differ by gender (p > 0.05) and were 0.77–5.04 mIU/L, 69.9–127.7 nmol/L, 3.84–6.20 pmol/L, 13.8–19.7 pmol/L, 1.8–51.2 µg/L, 0.9–1.2 TBI and 0.44–0.73 ng/mL, respectively. The RI for TT3 was different for males (1.49–2.53 nmol/L) and females (1.43–2.81 nmol/L), p = 0.021. A single cut-off for anti-TPO was established (<18 kIU/L). Differences in cut-offs for males and females were obtained for anti-Tg (<72 and <104 kIU/L, respectively) and TRAb (0.6 and 0.9 IU/L, respectively). Conclusions: This is the first study to determine RIs for thyroid function tests in Croatian adults on the Snibe analytical platform. The obtained results point out to the use of population- and immunoassay-specific RIs. For TT3, anti-Tg and TRAb gender-specific RIs should be considered. Full article

(This article belongs to the Special Issue Advances in Laboratory Analysis and Diagnostics)

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15 pages, 722 KB

Open AccessArticle

Evaluation of Interleukin-10, Vascular Endothelial Growth Factor Levels, and Bone Marrow Parameters in Multiple Myeloma Patients at Diagnosis and After Treatment

by Fulya Memis, Meryem Yalvac Kandefer, Sonay Aydin, Klara Dalva and Selami Kocak Toprak

Diagnostics 2025, 15(13), 1641; https://doi.org/10.3390/diagnostics15131641 - 27 Jun 2025

Cited by 1 | Viewed by 1737

Abstract

Background: Interleukin-10 (IL-10) and vascular endothelial growth factor (VEGF) are believed to possess a role in the pathophysiology of multiple myeloma (MM). We aimed to assess the significance of these parameters in the diagnosis, monitoring, and prognosis of the disease by examining them [...] Read more.

Background: Interleukin-10 (IL-10) and vascular endothelial growth factor (VEGF) are believed to possess a role in the pathophysiology of multiple myeloma (MM). We aimed to assess the significance of these parameters in the diagnosis, monitoring, and prognosis of the disease by examining them in patients at diagnosis and post-treatment and comparing the findings with those of healthy individuals. Methods: We conducted blood sampling from 35 patients diagnosed with MM at the time of diagnosis and from 15 of these patients post-treatment. We additionally assessed similar serum markers in a control group of 15 healthy individuals. Furthermore, we documented laboratory results, organ involvement, comorbidities, and CD27-CD81 levels assessed using flow cytometry in the bone marrow, along with treatments and patient responses. We also examined the quantity of cells collected during mobilization in patients who had autologous stem cell transplantation. Results: We found a positive correlation (p = 0.028/p = 0.035) between IL-10 and VEGF with the international staging score. In patients with renal involvement, IL-10 levels were higher and VEGF levels were lower than those without renal involvement (p = 0.011/p = 0.012). We showed that VEGF levels decreased significantly with treatment (p = 0.001). We found no statistically significant correlation between treatment responses and IL-10 and VEGF. The number of CD34 cells collected by mobilization showed a negative correlation with CD27 and a positive correlation with VEGF (p = 0.007/p = 0.032). Conclusions: Serum IL-10 level is associated with ISS and renal involvement in MM patients. There is a positive correlation between serum VEGF levels and the number of stem cells collected during mobilization. As CD27 expression increases, the number of stem cells collected in mobilization decreases. Full article

(This article belongs to the Special Issue Advances in Laboratory Analysis and Diagnostics)

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