Evaluation of Stability and Accuracy Compared to the Westergren Method of ESR Samples Analyzed at VES-MATIC 5.

The Erythrocyte Sedimentation Rate (ESR) is a diagnostic estimator of systemic inflammation as a reflection of acute phase proteins circulating in the blood. The purpose of this manuscript is to evaluate the blood stability at room temperature (RT) and at 4 °C to avoid ESR diagnostic errors, as well as the accuracy of the VES-MATIC 5 analyzer. The ESR stability evaluation at RT for 24 h (4 h "T1", 6 h "T2", 8 h "T3", 10 h "T4", 24 h "T5") and at 4 °C (24 h, 36 h, 48 h) was carried out using 635 total samples, starting with T0 (2 h of venipuncture). For method comparison, 164 patients were analyzed using VES-MATIC 5 and then the Westergren reference method. The sample at RT is established by a significant gradual decrease in correlation R = 0.99 (T0 vs. T1), R = 0.97 (T0 vs. T2), R = 0.92 (T0 vs. T3), R = 0.87 (T0 vs. T4), and R = 0.40 (T0 vs. T5). The stability at 4 °C after 24 h, 36 h, and 48 h showed a regression of R = 0.99, R = 0.97, and R = 0.95, respectively. Therefore, ESR measurements on RT samples beyond 6 h after collection cannot be carried out, but the ESR can be measured until 36 h for samples stored at 4 °C. Moreover, the VES-MATIC 5 accuracy performance compared to the Westergren method (R = 0.96) is confirmed.

Novel Biomarkers for Early Detection of Acute Kidney Injury and Prediction of Long-Term Kidney Function Decline after Partial Nephrectomy.

BACKGROUND: Identifying acute kidney injury (AKI) within few hours of onset is certainly helpful. However, early prediction of a long-term eGFR decline may be an even more important goal. Our aim was to identify and compare serum [creatinine, kineticGFR, cystatin C, neutrophil gelatinase-associated lipocalin (NGAL)] and urinary (NephroCheck, NGAL, proteinuria, albuminuria, acantocytes at urinary sediment) predictors of AKI that might efficiently predict long-term GFR decline after robotic Nephron-Spearing Surgery (rNSS). METHODS: Monocentric prospective observational study. Patients scheduled for rNSS for suspected localized Renal Cell Carcinoma from May 2017 to October 2017 were enrolled. Samples were collected preoperatively and postoperatively (timepoints: 4 h, 10 h, 24 h, 48 h), while kidney function was re-assessed up to 24 months. RESULTS: 38 patients were included; 16 (42%) developed clinical AKI. The eGFR decline at 24 months was more pronounced after postoperative AKI (-20.75 vs. -7.20, p < 0.0001). KineticGFR at 4 h (p = 0.008) and NephroCheck at 10 h (p = 0.001) were, at multivariable linear regression analysis, efficient predictors of post-operative AKI and long-term eGFR decline if compared to creatinine (R2 0.33 vs. 0.04). CONCLUSIONS: NephroCheck and kineticGFR have emerged as promising noninvasive, accurate, and early biomarkers of postoperative AKI and long-term GFR decline after rNSS. Combining NephroCheck and kineticGFR in clinical practice would allow to identify high risk of postoperative AKI and long-term GFR decline as early as 10 h after surgery.

Multicentric evaluation of the variability of digital morphology performances also respect to the reference methods by optical microscopy.

INTRODUCTION: Despite the important diagnostic role of peripheral blood morphology, cell classification is subjective. Automated image-processing systems (AIS) provide more accurate and objective morphological evaluation. The aims of this multicenter study were the evaluation of the intra and inter-laboratory variation between different AIS in cell pre-classification and after reclassification, compared with manual optical microscopy, the reference method. METHODS: Six peripheral blood samples were included in this study, for each sample, 70 May-Grunwald and Giemsa stained PB smears were prepared from each specimen and 10 slides were delivered to the seven laboratories involved. Smears were processed by both optical microscopy (OM) and AIS. In addition, the assessment times of both methods were recorded. RESULTS: Within-laboratory Reproducibility ranged between 4.76% and 153.78%; between-laboratory Precision ranged between 2.10% and 82.2%, while Total Imprecision ranged between 5.21% and 20.60%. The relative Bland Altman bias ranged between -0.01% and 20.60%. The mean of assessment times were 326 ± 110 s and 191 ± 68 s for AIS post reclassification and OM, respectively. CONCLUSIONS: AIS can be helpful when the number of cell counted are low and can give advantages in terms of efficiency, objectivity and time saving in the morphological analysis of blood cells. They can also help in the interpretation of some morphological features and can serve as learning and investigation tools.

Monocyte distribution width and the fighting action to neutralize sepsis (FANS) score for sepsis prediction in emergency department.

BACKGROUND AND AIMS: The aim of this study was to evaluate the diagnostic accuracy of sepsis markers and to develop a multiparametric score, using demographic and clinical variables as well as laboratory parameters to predict sepsis in patients admitted in the ED with suspected symptoms. MATERIALS AND METHODS: Patients with clinical presentation of suspected sepsis were enrolled in the ED of San Donato Hospital in Arezzo between September 2019 and May 2020. Anagraphic, anamnestic, clinical and laboratory data were collected for all subjects. PCT, MDW, WBC, MPV and BT were utilised to formulate FANS score. RESULTS: The AUC of the FANS score, PCT, MDW and CRP was 0.87, 0.80, 0.77 and 0.71, respectively, when used to predict sepsis in all 308 subjects. Instead, the AUC of the FANS (Fighting Action To Neutralize Sepsis) score, PCT, MDW and CRP was 0.93, 0.84, 0.83 and 0.77, respectively, when used to predict sepsis excluding subjects with infection (clinically classified as the Infections group). CONCLUSIONS: The results obtained with PCT, PCR and MDW confirm the results of these markers for the identification of sepsis obtained from other studies. The multiparametric approach, obtained from the statistical study of the parameters using binary logistic regression, identified those PCT, WBC, MPV, BT and MDW as the most significant and effective clinical classifiers for diagnosing sepsis.

Carriers of ADAMTS13 Rare Variants Are at High Risk of Life-Threatening COVID-19.

Thrombosis of small and large vessels is reported as a key player in COVID-19 severity. However, host genetic determinants of this susceptibility are still unclear. Congenital Thrombotic Thrombocytopenic Purpura is a severe autosomal recessive disorder characterized by uncleaved ultra-large vWF and thrombotic microangiopathy, frequently triggered by infections. Carriers are reported to be asymptomatic. Exome analysis of about 3000 SARS-CoV-2 infected subjects of different severities, belonging to the GEN-COVID cohort, revealed the specific role of vWF cleaving enzyme ADAMTS13 (A disintegrin-like and metalloprotease with thrombospondin type 1 motif, 13). We report here that ultra-rare variants in a heterozygous state lead to a rare form of COVID-19 characterized by hyper-inflammation signs, which segregates in families as an autosomal dominant disorder conditioned by SARS-CoV-2 infection, sex, and age. This has clinical relevance due to the availability of drugs such as Caplacizumab, which inhibits vWF-platelet interaction, and Crizanlizumab, which, by inhibiting P-selectin binding to its ligands, prevents leukocyte recruitment and platelet aggregation at the site of vascular damage.

The role of Monocyte Distribution Width (MDW) in the prognosis and monitoring of COVID-19 patients.

The new parameter derived from the standard deviation of the monocyte distribution width (MDW) has shown a good diagnostic efficacy in COVID-19 patients. In this study, we propose MDW as a prognostic and monitoring parameter in patients with severe forms of COVID-19. Sixty SARS-CoV-2-positive patients admitted to the San Donato Hospital in Arezzo were enrolled. A blood sample taken to measure the complete blood count was used for the determination of MDW using a UniCel DxH 900 instrument (Beckman Coulter). For each patient, a mean of 6 ± 2 measurements of MDW were taken. The difference between the last and first MDW results was reported as the ΔMDW variable. The ΔMDW and age were significantly correlated to the outcome. In non-survivors patients, the difference in the mean of the MDW between the first and other points was not significant, while in survivors, the first point was higher than the other points (p < 0.005), with the exception of the mean of the second point (p-value = NS). The ΔMDW area under the curve (AUC) was 0.84, and with a cut-off lower than 0.00 the sensitivity and specificity were 88% and 81%, respectively. The most important result of this study is the ΔMDW calculated on the basis of the difference between the first and third measurement, after approximately the 5-7th day of hospitalisation. A ΔMDW less than one was indicative of an unfavourable prognosis. The data reported suggest that MDW could be used to support monitoring and surveillance, alongside other tests such as procalcitonin, in critically ill patients in the ICU.

Isolated case of platelet satellitism around white blood cells and phagocytosis by neutrophils and monocytes.

INTRODUCTION: Rarely, platelets can interact with other blood elements, forming platelet aggregates. This paper presents an isolated case of platelet satellitism around neutrophils, lymphocytes and monocytes with platelet phagocytosis by both neutrophils and monocytes. CASE PRESENTATION: The subject was an 89-year-old woman with breast cancer on anti-estrogenic hormone cancer therapy. Whole blood sample collected in a tube with EDTA (Ethylenediaminetetra-acetic acid) anticoagulant was analysed within 4 hours, using an XN haematology analyser (Sysmex). The CBC (complete blood count) presented the following results: WBC (White blood cell) 4.0 x 109/L, RBC (Red blood cell) 3.58 x 1012/L, haemoglobin 116 g/L, haematocrit 34.9%, MCV (Mean corpuscular volume) 97.5 fL, MCH (Mean corpuscular haemoglobin) 32.5 pg, MCHC (Mean corpuscular haemoglobin concentration) 33.2 g /dL, RDW (Red blood cell distribution width) 14.6% and PLT (Platelet) 136. CONCLUSION: This manuscript describes the platelet satellitism around neutrophils, lymphocytes and monocytes and the interesting, very rare and singular phenomenon of platelet phagocytosis by not only neutrophils but also monocytes.

Multicentric evaluation of analytical performances digital morphology with respect to the reference methods by manual optical microscopy.

AIMS: Optical microscopic (OM) evaluation of peripheral blood (PB) cells is still a crucial step of the laboratory haematological workflow. The morphological cell analysis is time-consuming and expensive and it requires skilled operator. To address these challenges, automated image-processing systems, as digital morphology (DM), were developed in the last few years. The aim of this multicentre study, performed according to international guidelines, is to verify the analytical performance of DM compared with manual OM, the reference method. METHODS: Four hundred and ninety PB samples were evaluated. For each sample, two May Grunwald-stained and Giemsa-stained smears were performed and the morphological evaluation of cells was analysed with both DM and OM. In addition, the assessment times of both methods were recorded. RESULTS: Comparison of DM versus OM methods was assessed with Passing-Bablok and Deming fit regression analysis: slopes ranged between 0.17 for atypical, reactive lymphocytes and plasma cells (LY(AT)) and 1.24 for basophils, and the intercepts ranged between -0.09 for blasts and 0.40 for LY(AT). The Bland-Altman bias ranged between -6.5% for eosinophils and 21.8% for meta-myemielocytes. The diagnostic agreement between the two methods was 0.98. The mean of assessment times were 150 s and 250 s for DM and OM, respectively. CONCLUSION: DM shows excellent performance. Approximately only 1.6% of PB smears need the OM revision, giving advantages in terms of efficiency, standardisation and assessment time of morphological analysis of the cells. The findings of this study may provide useful information regarding the use of DM to improve the haematological workflow.

Improvement in estimation of time since death by albumin and potassium concentrations in vitreous humor.

The measurement of the potassium concentration (K+) in vitreous humor (VH) has been a well-known adjunct for the estimation of the post-mortem interval (PMI) since the early 1960s. For years, however, many authors have been using other biochemical markers in an attempt to improve predictions. In this paper we confirm the role of K+ in the determination of the PMI adopting a linear regression model and we investigate whether other biochemical markers could improve the model through a multiple regression analysis. Additionally, the research aims to confirm the data of the analytes of interest among different techniques and instrumentations. We deemed this as an important issue because a primary concern in the literature is that automated analytical methods are often calibrated and, for the most part, validated for serum or urine analysis. Our results confirmed the well-established role of K+ as well as highlighted Albumin as a novel marker to be considered for further improvement of prediction models, especially since 72 h after death.

Comparison of serologic and molecular SARS-CoV 2 results in a large cohort in Southern Tuscany demonstrates a role for serologic testing to increase diagnostic sensitivity.

BACKGROUND: Since February 2020, Italian hospitals registered COVID-19 (COronaVIrus Disease 19) cases more often than the rest of the Europe. During this epidemic, health authorities requested swab tests, while seeking new patient paths. METHODS: A dual laboratory approach was evaluated, consisting of patient care reports for viral RNA detection on swabs and rapid serological tests in 516 patients (192 symptomatic or paucisymptomatic and 324 asymptomatic). RESULTS: We found the molecular positive fraction equal to 12% (23/192) among symptomatic/paucisymptomatic (S/P) and 15.4% (50/324) in asymptomatic (As) sets. Among subsets, we observed serologically positive results, corresponding to 35% (8/23) for S/P and 38% (19/50) for As. Among molecular negative cases, we detected specific Immunoglobulin G or M (Ig G or Ig M) positivity in the S/P cohort equal to 6.6% (11/167) and 6% (15/246) in As cases. For indeterminate molecular results, we found S/P serological positivity equal to 100% (1/1) and 54% (13/24) in As patients. We found higher (p < 0.05) seropositivity in older patients (n = 8) among symptomatic and positives for viral RNA (n.23). CONCLUSIONS: It has been observed that a dual approach of serological and molecular tests detects a higher absolute number of disease cases in a pandemic context,which could improve monitoring and health surveillance efficacy. The age-related seropositivity frequency in this study, if confirmed, could enhance the validity of serological tests, especially in older patients.In these subjects, molecular positivity accompanied by serological positivity (distinct for M and G immunoglobulins) should help determine disease status and support decisions related to patient management.

Elevated monocyte distribution width in COVID-19 patients: The contribution of the novel sepsis indicator.

INTRODUCTION: Interesting results regarding the contribution of MDW (Monocyte Distribution Width) in the Infectious Disease Unit have been reported. An observational study is ongoing at San Donato Hospital with the aim to evaluate the contribution of MDW in the diagnostic pathway in adult patients entering in the ED setting and tested for SARS-CoV-2. MATERIAL AND METHOD: COVID-19 symptomatic and paucisymptomatic patients presenting to ED (Emergency Department), have been enrolled consecutively. Whole blood venous samples have been collected on K2 EDTA for MDW determination, at the same time a nasopharyngeal swab for SARS-CoV-2 RNA detection have been collected. RESULTS: One hundred six patients were negative for SARS-CoV-2 with MDW mean value of 20.3 ± 3.3, while forty-one were positive for SARS-CoV-2 with higher MDW mean value of 27.3 ± 4.9 (P < 0.005). The ROC curve analysis has been evaluated showing MDW AUC of 0.91. Finally twenty-three patients hospitalized in high-intensity care unit showed an MDW value higher than the eighteen patients presenting few symptoms [28.8 ± 5.3 vs 25.4 ± 3.6 respectively, P < 0.05]. DISCUSSION: Monocytic population, in Covid19 disease, are the first elements of innate immunity to be involved, these changes are the basis of the modification of the MDW, with evident efficacy in term of sensitivity, particularly in the studied Covid19 patients. Moreover the patients hospitalized in high-intensity care unit showed significantly elevated MDW respects to middle or low symptomatic one, suggest including this parameter as prognostic marker or of therapy efficacy, integrated with other laboratory findings.

Multicenter evaluation of analytical performances of platelet counts and platelet parameters: Carryover, precision, and stability.

INTRODUCTION: The correctness of the results of automated platelet analysis is still highly debated. The aim of this multicenter study, conducted according to international guidelines, was to verify the analytical performance of nine different types of hematology analyzers (HAs) in the automated platelet analysis. METHODS: Four hundred eighty-six peripheral blood samples (PB), collected in K3 EDTA tubes, were analyzed by ABX Pentra, ADVIA2120i, BC-6800, BC-6800 Plus, Cell-DYN Sapphire, DxH800, XE-2100, XE-5000, XN-20 with PLT-F App. Within-run imprecision and between-run imprecision were carried out using PB and material control, respectively. The carryover, low limit of quantification (LoQ), and the PB stability were evaluated. RESULTS: The carryover was absent for all HAs. The LoQ of PLT ranged between 2.0 (Cell-Dyn Sapphire) and 25.0 × 109 /L (ADVIA 2120i), while immature platelet fraction (IPF) ranged between 1.0 (XN-20) and 12.0 × 109 /L (XE-5000). The imprecision (%CV) increases as the platelet count decreases. No HAs showed desirable CVAPS for PLT counts less than 50.0 × 109 /L, with the exception of Cell-DYN Sapphire (CV 3.0% with PLT-O mean value of 26.7 × 109 /L), XN-20 (CV 2.4% with PLT-F mean value of 21.5 × 109 /L), and BC-6800 Plus (CV 1.9% with PLT-O mean value of 26.5 × 109 /L). The sample stability ranged between under two hours for MPV by ADVIA2120i and 8 hours for other PLT parameters and HAs. CONCLUSION: The findings of this study may provide useful information regarding carryover, precision, and stability of platelet counts and parameters, especially in thrombocytopenic samples. Moreover, the stability of sample for platelet analysis is conditioned by the HA and by temperature and storage time.

Seminal but not Serum Levels of Holotranscobalamin are Altered in Morbid Obesity and Correlate with Semen Quality: A Pilot Single Centre Study.

Vitamin B12 (cobalamin) is an essential cofactor in the one-carbon metabolism. One-carbon metabolism is a set of complex biochemical reactions, through which methyl groups are utilised or generated, and thus plays a vital role to many cellular functions in humans. Low levels of cobalamin have been associated to metabolic/reproductive pathologies. However, cobalamin status has never been investigated in morbid obesity in relation with the reduced semen quality. We analysed the cross-sectional data of 47-morbidly-obese and 21 lean men at Careggi University Hospital and evaluated total cobalamin (CBL) and holotranscobalamin (the active form of B12; holoTC) levels in serum and semen. Both seminal and serum concentrations of holoTC and CBL were lower in morbidly obese compared to lean men, although the difference did not reach any statistical significance for serum holoTC. Seminal CBL and holoTC were significantly higher than serum levels in both groups. Significant positive correlations were observed between seminal holoTC and total sperm motility (r = 0.394, p = 0.012), sperm concentration (r = 0.401, p = 0.009), total sperm number (r = 0.343, p = 0.028), and negative correlation with semen pH (r = -0.535, p = 0.0001). ROC analysis supported seminal holoTC as the best predictor of sperm number (AUC = 0.769 ± 0.08, p = 0.006). Our findings suggest that seminal rather than serum levels of holoTC may represent a good marker of semen quality in morbidly obese subjects.