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Valaperta R.,Research Laboratories Molecular Biology | Sansone V.,Stroke Unit and Center for Neuromuscular Disease | Lombardi F.,Research Laboratories Molecular Biology | Verdelli C.,Research Laboratories Molecular Biology | And 6 more authors.
BioMed Research International | Year: 2013

The expansion of the specific trinucleotide sequence, [CTG], is the molecular pathological mechanism responsible for the clinical manifestations of DM1. Many studies have described different molecular genetic techniques to detect DM1, but as yet there is no data on the analytical performances of techniques used so far in this disease. We therefore developed and validated a molecular method, "Myotonic Dystrophy SB kit," to better characterize our DM1 population. 113 patients were examined: 20 DM1-positive, 11 DM1/DM2-negative, and13 DM1-negative/DM2-positive, who had a previous molecular diagnosis, while 69 were new cases. This assay correctly identified 113/113 patients, and all were confirmed by different homemade assays. Comparative analysis revealed that the sensitivity and the specificity of the new kit were very high (>99%). Same results were obtained using several extraction procedures and different concentrations of DNA. The distribution of pathologic alleles showed a prevalence of the "classical" form, while of the 96 nonexpanded alleles 19 different allelic types were observed. Cardiac and neuromuscular parameters were used to clinically characterize our patients and support the new genetic analysis. Our findings suggest that this assay appears to be a very robust and reliable molecular test, showing high reproducibility and giving an unambiguous interpretation of results. © 2013 Rea Valaperta et al.


PubMed | Abbott Laboratories, Chioggia and Service of Laboratory Medicine
Type: | Journal: Blood transfusion = Trasfusione del sangue | Year: 2014

Blood donors positive only for anti-HBc may have a resolved hepatitis B virus (HBV) infection, low grade chronic infection or infection with variant strains of HBV. We aimed to assess the significance of this serological pattern after hepatitis B vaccination in such cases.Twenty-four anti-HBc only blood donors were vaccinated with the Engerix HBV vaccine and a serological and virological evaluation was performed before HBV vaccination and 7-10 days after each dose. Subjects were classified as non-responders if their anti-HBs levels stayed below 10 IU/L after full vaccination, while the response was considered secondary (anamnestic) if anti-HBs levels rose over 10 IU/L after the first vaccine dose, and primary if anti-HBs levels rose over 10 IU/L only after the second or third vaccine dose.Of the 21 fully evaluable donors, six had no response, eight showed a primary response and seven had an anamnestic response. One non-responder had transient positivity for HBV-DNA at low levels (12 IU/mL) with persistent negativity for HBsAg.Anti-HBc-only positive blood donors are a heterogeneous population including HBV nave subjects with a likely false-positive anti-HBc reactivity, subjects with a resolved HBV infection, and subjects with persistent low-level HBV replication. The analysis of the anti-HBs response after a dose of HBV vaccine may help to distinguish among the different causes of the isolated anti-HBc positivity, thereby enabling proper counselling and potential readmission to blood donation.


Gessoni G.,Transfusional Service | Gessoni G.,Service of Laboratory Medicine | Valverde S.,Service of Laboratory Medicine | Canistro R.,Onco Hematology Unit | And 3 more authors.
Biochimica Clinica | Year: 2013

Acquired hemophilia A (AHA) is a rare, but often life-threatening hemorrhagic disorder characterized by antibodies directed against coagulation factor VIII. We report clinical and laboratory investigations of two cases with AHA observed in our hospital. These patients were two elderly women (73 and 62 years old), who presented with subcutaneous bleeding, intramuscular hematoma and a prolonged activated partial thromboplastin time (aPTT). On the basis of these findings as well as decreased factor VIII activities and the presence of factor VIII inhibitors, we made a diagnosis of AHA. Both patients were referred to a specialized hospital for treatment. The diagnosis of AHA should be considered in any elderly patient who presents with bleeding and prolonged aPTT. Moreover, the coexistence of a series of underlying diseases associated with AHA should be always searched for.


PubMed | University of Pavia, Service of Laboratory Medicine, Research Laboratories, Clinical Cardiology Unit and CCU and 3 more.
Type: | Journal: Clinica chimica acta; international journal of clinical chemistry | Year: 2016

Myotonic dystrophy (DM) is a genetic disorder caused by nucleotide repeats expansion. Sudden death represents the main cause of mortality in DM patients. Here, we investigated the relationship between serum cardiac biomarkers with clinical parameters in DM patients.Case-control study included 59 DM patients and 22 healthy controls. An additional group of 62 controls with similar cardiac defects to DM were enrolled.NT-proBNP, hs-cTnT and CK levels were significantly increased in DM patients compared to healthy subjects (p=0.0008, p<0.0001, p<0.0001). Also, hs-cTnT levels were significantly higher in DM compared to control group with cardiac defects (p=0.0003). Positive correlation was found between hs-cTnT and hs-cTnI in both DM patients and controls (p=0.019, p=0.002). Independently from the age, the risk of DM disease was positively related to an increase in hs-cTnT (p=0.03). On the contrary, the risk of DM was not related to hs-cTnI, but was evidenced a role of PR interval (p=0.03) and CK (p=0.08).The levels of hs-cTnT were significantly higher in DM patients. Analysis, with anti-cTnT, shows that this increase might be linked to heart problems. This last finding suggests that hs-cTnT might represent a helpful serum biomarker to predict cardiac risk in DM disease.


Trape J.,Service of Laboratory Medicine | Molina R.,Hospital Clinic Of Barcelona | Sant F.,Service of Pathology | Montesinos J.,Service of Medical Oncology | And 11 more authors.
Tumor Biology | Year: 2012

The utility of tumour markers (TM) in the differential diagnosis of cancer in serous effusion (fluid effusion (FE)) has been the subject of controversy. The aim of this study was to prospectively validate our previous study and to assess whether the addition of adenosine deaminase (ADA), C-reactive protein (CRP) or percentage of polymorphonuclear cells (%PN) allows the identification of false positives. In this study, carcinoembryonic antigen, cancer antigen 15-3, cancer antigen 19-9, ADA, CRP and %PN in FE were determined in 347 patients with 391 effusions. Effusions were considered as malignant effusion when at least one TM in serum exceeded the cutoff and the ratio FE/S was higher than 1.2. Also, cases with values of ADA, CRP and %PN above the established cutoffs in serous effusion were considered as potential false positives. The combined sensitivity and specificity of the three TM was 76.2 % (95 % confidence intervals (CI) 67.8-83.3 %) and 97.0 % (95 % CI 94.1-98.7), respectively. Subanalysis of the 318 cases with previous criteria and negative ADA, CRP and %PN obtained sensitivities of 78.4 % (95 % CI 69.4-85.6) and a specificity of 100 % (95 % CI 98.2-100). The results obtained validate our previous study and are improved with the addition of ADA, CRP and %PN. TM in serous effusions and serum could be useful for the diagnostic assessment of patients with serous effusions. © 2012 International Society of Oncology and BioMarkers (ISOBM).


Renna L.V.,University of Milan | Cardani R.,Laboratory of Muscle Histopathology and Molecular Biology | Botta A.,University of Rome Tor Vergata | Rossi G.,University of Rome Tor Vergata | And 5 more authors.
European Journal of Histochemistry | Year: 2014

Myotonic dystrophy type 1 (DM1) and type 2 (DM2) are multisystemic disorders linked to two different genetic loci and characterized by several features including myotonia, muscle weakness and atrophy, cardiac dysfunctions, cataracts and insulin-resistance. In both forms, expanded nucleotide sequences cause the accumulation of mutant transcripts in the nucleus deregulating the activity of some RNAbinding proteins and providing an explanation for the multisystemic phenotype of DM patients. However this pathogenetic mechanism does not explain some histopathological features of DM skeletal muscle like muscle atrophy. It has been observed that DM muscle shares similarities with the ageing muscle, where the progressive muscle weakness and atrophy is accompanied by a lower regenerative capacity possibly due to the failure in satellite cells activation. The aim of our study is to investigate if DM2 satellite cell derived myoblasts exhibit a premature senescence as reported for DM1 and if alterations in their proliferation potential and differentiation capabilities might contribute to some of the histopathological features observed in DM2 muscles. Our results indicate that DM myoblasts have lower proliferative capability than control myoblasts and reach in vitro senescence earlier than controls. Differentely from DM1, the p16 pathway is not responsible for the premature growth arrest observed in DM2 myoblasts which stop dividing with telomeres shorter than controls. During in vitro senescence, a progressive decrease in fusion index is observable in both DM and control myotubes with no significant differences between groups. Moreover, myotubes obtained from senescent myoblasts appear to be smaller than those from young myoblasts. Taken together, our data indicate a possible role of DM2 premature myoblast senescence in skeletal muscle histopathological alterations i.e., dystrophic changes and type 2 fibre atrophy. © L.V. Renna et al., 2014.


Gessoni G.,Service of Laboratory Medicine | Valverde S.,Service of Laboratory Medicine | Manoni F.,Service of Laboratory Medicine
Clinica Chimica Acta | Year: 2012

Background: In this study, we evaluated the GeneXpert HemosIL Factor II and Factor V assay, an innovative assay for the detection of Factor V Leiden (FVL) and prothrombin G20210A mutation (GPRO). Patients and methods: We evaluated 132 patients that were previously classified (with a concordant result) using two commercial real-time PCR assays supplied, by Applied Biosystems and Roche Molecular Biochemicals. The cohort comprised 75 normal subjects, 10 FVL homozygous, 35 FVL heterozygous, 7 GPRO heterozygous, 2 GPRO homozygous and 3 double heterozygous FVL and GPRO subjects. All of the samples were evaluated using the GeneXpert HemosIL Factor II and Factor V assay. Results: All of the samples were correctly identified using the GeneXpert HemosIL Factor II and Factor V assay; therefore, in this patient series, the specificity and sensitivity of the test under evaluation was 1.00. Discussion: We have shown that the GeneXpert HemosIL Factor II and Factor V assay, a rapid fully automated assay, can accurately characterise the presence of FV G1691A and FII G20210A polymorphisms with specificity and sensitivity that are comparable to other current real-time PCR-based methods. The theoretical advantages of such an assay include improved standardisation across varying healthcare environments, more thorough sample manipulation and reduced human error. © 2012 Elsevier B.V.


Valverde S.,Service of Laboratory Medicine | Penzo L.,Service of Laboratory Medicine | Gessoni G.,Service of Laboratory Medicine
Biochimica Clinica | Year: 2013

In this study we evaluated efficiency and quality in our laboratory after the implementation of total laboratory automation for clinical chemistry and immunochemistry assays. Thermo EnGen pre-analytical automation with a track system connecting two Vitros Fusion 5.1 and two Tosoh AIA-2000 analyzers were implemented. For Vitros Fusion 5.1, CVs were <8% (within-run) and <10% (inter-assay), except for measurement of anti-streptolysin O titre (CV 22.7%-27.7%). For Tosoh AIA-2000 analyzers, within-run CVs were <10% and between-run CVs <11%. No drift effects were observed, neither there was any carry-over. When evaluating the functionality of the whole automation system, we observed a turnaround time (TAT) 90% <10 min for tubes needing check-in and exit only. For tubes needing check-in and centrifugation - exit cycle, TAT 90% was <30 min. Result availability for clinical chemistry and/or immunochemistry on-line analyzers showed a TAT 90% <120 min. The adopted automation system effectively reduced the labor associated with specimen processing, possibly decreasing the number of laboratory errors that occur with specimen sorting, labeling and aliquoting, and improved the integrity of specimen handling throughout steps of specimen processing.


Manoni F.,Service of Laboratory Medicine | Tinello A.,Service of Laboratory Medicine | Fornasiero L.,Service of Laboratory Medicine | Hoffer P.,Service of Laboratory Medicine | And 3 more authors.
Clinical Chemistry and Laboratory Medicine | Year: 2010

Background: The study of urine particles plays a key role in the diagnosis of kidney diseases. In this study, the authors evaluated the correlation between the UF-1000i and quantitative manual microscopy. Methods: A total of 214 untreated urine samples were studied using the Sysmex UF-1000i and compared with results obtained from quantitative manual microscopy using the Fuchs-Rosenthal counting chamber. Results: Using Pearson statistics, we observed satisfactory correlation between the UF-1000i and quantitative microscopy: for red blood cells (RBCs) r was 0.98, for white blood cells (WBCs) r was 1.00, for epithelial cells (EC) r was 0.96, and for casts r was 0.69. Using linear regression statistics, we also observed satisfactory correlation between the UF-1000i and quantitative microscopy: for RBCs R2 was 0.95, for WBCs R2 was 0.99, for EC R2 was 0.92, and for casts R2 was 0.48. Conclusions: In our experience, automated urine particle analysis performed using the Sysmex UF-1000i analyzer is sufficiently precise and improves the workflow in a routine laboratory. Precision was satisfactory and concordance with the reference method is good for RBC, WBC and EC; for casts microscopic observation is required for flagged samples to discriminate hyaline from pathologic casts. © 2010 by Walter de Gruyter Berlin New York.


PubMed | Service of Laboratory Medicine
Type: Journal Article | Journal: Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine | Year: 2012

The utility of tumour markers (TM) in the differential diagnosis of cancer in serous effusion (fluid effusion (FE)) has been the subject of controversy. The aim of this study was to prospectively validate our previous study and to assess whether the addition of adenosine deaminase (ADA), C-reactive protein (CRP) or percentage of polymorphonuclear cells (%PN) allows the identification of false positives. In this study, carcinoembryonic antigen, cancer antigen 15-3, cancer antigen 19-9, ADA, CRP and %PN in FE were determined in 347 patients with 391 effusions. Effusions were considered as malignant effusion when at least one TM in serum exceeded the cutoff and the ratio FE/S was higher than 1.2. Also, cases with values of ADA, CRP and %PN above the established cutoffs in serous effusion were considered as potential false positives. The combined sensitivity and specificity of the three TM was 76.2 % (95 % confidence intervals (CI) 67.8-83.3 %) and 97.0 % (95 % CI 94.1-98.7), respectively. Subanalysis of the 318 cases with previous criteria and negative ADA, CRP and %PN obtained sensitivities of 78.4 % (95 % CI 69.4-85.6) and a specificity of 100 % (95 % CI 98.2-100). The results obtained validate our previous study and are improved with the addition of ADA, CRP and %PN. TM in serous effusions and serum could be useful for the diagnostic assessment of patients with serous effusions.

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