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1.
Langenbecks Arch Surg ; 409(1): 285, 2024 Sep 20.
Artículo en Inglés | MEDLINE | ID: mdl-39302485

RESUMEN

PURPOSE: In this study, we analyse the possibility to omit pre-incision PTH measurement since we routinely measure it at the time of pre-surgery ambulatory admission. METHODS: A total of 435 patients were enrolled. All patients with pHPT included underwent pre-surgical PTH level assessment as part of the pre-admission preparation to surgery. Intraoperative PTH was routinely assessed after induction of the anaesthesia (pre-incision PTH) and 15 min after resection of the enlarged gland(s) (post-excision PTH). Moreover, calcium and PTH levels were routinely assessed on the first postoperative day. Cure was defined as an intraoperative drop of > 50% or into normal range on first post-operative day. RESULTS: The median value of the preoperative and pre-incision PTH were both 127 pg/ml (p = ns). Thirty-two patients (7.3%) exhibited a not appropriate drop of post-excision PTH level. Nevertheless, nineteen of them (59.3%) showed a satisfying PTH drop on 1st POD. Ten patients (2.3%) experienced a persistent disease with six achieving cure through reoperation. Additionally, three patients (0.6%) showed normalization of calcium and PTH values during the follow-up. Three patients, apparently deemed cured after an adequate PTH-drop on the day of surgery, showed persistence. Cure rate at primary surgery was 98.4%. Accuracy of our simplified protocol is 99.3%. CONCLUSION: Pre-incision PTH is not superior to preoperative PTH blood test and can be omitted without compromising the sensitivity of cure prediction. One blood sample 15 min after resection, along with the postoperative PTH value on the day after surgery, is sufficient to predict the surgical outcome bearing the cost of a very low reoperation rate.

2.
BMC Infect Dis ; 22(1): 291, 2022 Mar 26.
Artículo en Inglés | MEDLINE | ID: mdl-35346089

RESUMEN

BACKGROUND: The aim of our study was to assess the impact the impact of gender and age on reactogenicity to three COVID-19 vaccine products: Biontech/Pfizer (BNT162b2), Moderna (mRNA-1273) and AstraZeneca (ChAdOx). Additional analyses focused on the reduction in working capacity after vaccination and the influence of the time of day when vaccines were administered. METHODS: We conducted a survey on COVID-19 vaccinations and eventual reactions among 73,000 employees of 89 hospitals of the Helios Group. On May 19th, 2021 all employees received an email, inviting all employees who received at least 1 dose of a COVID-19 to participate using an attached link. Additionally, the invitation was posted in the group's intranet page. Participation was voluntary and non-traceable. The survey was closed on June 21st, 2021. RESULTS: 8375 participants reported on 16,727 vaccinations. Reactogenicity was reported after 74.6% of COVID-19 vaccinations. After 23.0% vaccinations the capacity to work was affected. ChAdOx induced impairing reactogenicity mainly after the prime vaccination (70.5%), while mRNA-1273 led to more pronounced reactions after the second dose (71.6%). Heterologous prime-booster vaccinations with ChAdOx followed by either mRNA-1273 or BNT162b2 were associated with the highest risk for impairment (81.4%). Multivariable analyses identified the factors older age, male gender and vaccine BNT162b as independently associated with lower odds ratio for both, impairing reactogenicity and incapacity to work. In the comparison of vaccine schedules, the heterologous combination ChAdOx + BNT162b or mRNA-1273 was associated with the highest and the homologue prime-booster vaccination with BNT162b with the lowest odds ratios. The time of vaccination had no significant influence. CONCLUSIONS: Around 75% of the COVID-19 vaccinations led to reactogenicity and nearly 25% of them led to one or more days of work loss. Major risk factors were female gender, younger age and the administration of a vaccine other than BNT162b2. When vaccinating a large part of a workforce against COVID-19, especially in professions with a higher proportion of young and women such as health care, employers and employees must be prepared for a noticeable amount of absenteeism. Assuming vaccine effectiveness to be equivalent across the vaccine combinations, to minimize reactogenicity, employees at risk should receive a homologous prime-booster immunisation with BNT162b2. TRIAL REGISTRATION: The study was approved by the Ethic Committee of the Aerztekammer Berlin on May 27th, 2021 (Eth-37/21) and registered in the German Clinical Trials Register (DRKS 00025745). The study was supported by the Helios research grant HCRI-ID 2021-0272.


Asunto(s)
Vacunas contra la COVID-19 , COVID-19 , Vacuna BNT162 , COVID-19/prevención & control , Vacunas contra la COVID-19/efectos adversos , Vacuna contra Difteria, Tétanos y Tos Ferina , Femenino , Personal de Salud , Humanos , Masculino , Vacunación
3.
Clin Chem Lab Med ; 59(2): 301-306, 2020 05 08.
Artículo en Inglés | MEDLINE | ID: mdl-32383687

RESUMEN

Objectives: Quality management for clinical laboratories requires the establishment of internal procedures including standard operating procedures (SOPs), internal quality control (QC), validation of test results and quality assessment. External quality assessment (EQA) and alternativeassessment procedures (AAPs) are part of the quality hierarchy required for diagnostic testing. The International Organization for Standardization (ISO) document with requirements for conformance ISO 15189 and the Clinical and Laboratory Standards Institute document (CLSI) QMS24 require participation in EQA schemes and AAPs where applicable. The purpose of this study was to perform a global survey of EQA and AAPs for key procedures in molecular diagnostic laboratories. Methods: The Committee for Molecular Diagnostics of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC C-MD) conducted a survey of international molecular laboratories that covered specific topics of molecular diagnostic services as well as methods for EQA and AAPs. The survey addressed the following aspects: (1) usage of laboratory-developed test (LDT), (2) participation in EQA schemes and (3) performance of AAPs. Results: A total of 93 responses from laboratories located in Asia, Europe, the Middle East, North America and South America were received. The majority of the participating laboratories (65.9%) use LDTs and 81.3% stated that it is mandatory for them to participate in EQA programs, while 22% of the laboratories reported not performing AAPs. Thirty-one percent of the laboratories use EQAs for fewer than 50.0% of their reported parameters/analytes. Conclusions: While the majority of laboratories perform EQA and AAPs to improve their quality in molecular diagnostics, the amount of AAPs as quality procedures differs within the laboratories. Further surveys are necessary to clarify the existing needs in additional EQAs and standardized AAPs. The survey will also guide future efforts of the IFCC C-MD for identifying quality practices in need to improve harmonization and standardization within molecular diagnostics.


Asunto(s)
Laboratorios/normas , Patología Molecular/métodos , Garantía de la Calidad de Atención de Salud/normas , Control de Calidad , Técnicas de Laboratorio Clínico , Técnicas y Procedimientos Diagnósticos , Humanos , Estándares de Referencia , Encuestas y Cuestionarios
4.
Palliat Med ; 34(6): 776-783, 2020 06.
Artículo en Inglés | MEDLINE | ID: mdl-32186249

RESUMEN

BACKGROUND: Multidrug-resistant organisms are a growing challenge and burden to patient care. To date, there are only data concerning the prevalence of methicillin-resistant Staphylococcus aureus infections. Thus, numbers of other multidrug-resistant organisms can only be extrapolated and inferred from more or less comparable cohorts. AIM: To evaluate the prevalence of multidrug-resistant organisms on palliative care in-patients. DESIGN: A prospective cohort analysis. SETTING/PARTICIPANTS: A University Hospital-bound palliative care unit, in which all patients admitted to the unit were screened for inclusion. RESULTS: In total, 304 patients were included in this study. The prevalence for methicillin-resistant Staphylococcus aureus of 5.2% (95% confidence interval: 2.9%-8.4%), for vancomycin-resistant Enterococcus faecium of 10.5% (95% confidence interval: 7.2%-14.8%), for Ciprofloxacin-resistant-extended spectrum beta-lactamases isolates of 5.8% (95% confidence interval: 3.4%-9.3%) and Ciprofloxacin-resistant Carbapenem-resistant Gram-negative bacteria of 0.3% (95% confidence interval: 0%-1.3%) was calculated. Except for methicillin-resistant Staphylococcus aureus, patients carrying a multidrug-resistant organism had a significant longer duration of hospitalization. Median length of stay was 12 days (interquartile range: 14.5, no multidrug-resistant organisms), 14.5 days (interquartile range: 15, methicillin-resistant Staphylococcus aureus), 21 days (interquartile range: 16.5, vancomycin-resistant enterococci), 22 days (interquartile range: 20.75, Ciprofloxacin-resistant-extended spectrum beta-lactamases) and 32 days (interquartile range: 22.00) for patients carrying two organisms. CONCLUSION: There is a high prevalence of all multidrug-resistant organisms within the hospitalized palliative care patients. However, the multidrug-resistant organisms do not seem to impact the survival within this cohort. Further studies should evaluate additional end-points, for example, quality of life, which are of special interest in this cohort.


Asunto(s)
Bacterias , Farmacorresistencia Bacteriana , Hospitales , Cuidados Paliativos , Bacterias/efectos de los fármacos , Bacterias/aislamiento & purificación , Infecciones Bacterianas/epidemiología , Infecciones Bacterianas/microbiología , Estudios de Cohortes , Hospitales/estadística & datos numéricos , Humanos , Tiempo de Internación , Cuidados Paliativos/estadística & datos numéricos , Prevalencia , Estudios Prospectivos , Calidad de Vida , Análisis de Supervivencia
5.
Clin Chem Lab Med ; 57(8): 1271-1279, 2019 07 26.
Artículo en Inglés | MEDLINE | ID: mdl-30763260

RESUMEN

Background The increasing number of multi-drug resistant (MDR) bacteria provides enormous challenges for choosing an appropriate antibiotic therapy in the early phase of sepsis. While bacterial identification has been greatly accelerated by the introduction of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), the antibiotic susceptibility testing (AST) remains time-consuming. Here, we present a rapid susceptibility testing method for testing Gram-negative bacteria, exemplarily validated for Escherichia coli and Klebsiella spp. Methods Gram-negative isolates (E. coli and Klebsiella spp.) were either taken as single colonies from agar plates (n=136) or directly extracted and identified from positive blood cultures (n=42) using MALDI-TOF MS. Bacteria were incubated in glucose-supplemented Luria broths (LBs) each containing one antibiotic (ceftazidime, piperacillin, imipenem and ciprofloxacin), routinely used to classify Gram-negative bacteria in Germany. To determine susceptibility the dynamics of glucose utilization in bacterial suspensions were quantitatively measured in the presence or absence of antibiotics designated liquid-AST (L-AST). Results The L-AST can be run on clinical-chemistry analyzers and integrated into laboratory routines. It yields critical resistance information within 90-150 min downstream of a MS-based identification. The results showed a high concordance with routine susceptibility testing, with less than 1% very major errors (VME) and 3.51% major errors (ME) for 178 assessed isolates. Analysis of turnaround time (TAT) for 42 clinical samples indicated that L-AST results could be obtained 34 h earlier than the routine results. Conclusions As exemplified for E. coli and Klebsiella spp., L-AST provides substantial acceleration of susceptibility testing following MALDI-TOF MS identification. The assay is a simple and low-cost method that can be integrated into clinical laboratory to allow for 24/7 AST. This approach could improve antibiotic therapy.


Asunto(s)
Pruebas de Química Clínica , Escherichia coli/aislamiento & purificación , Glucosa/análisis , Glucosa/metabolismo , Klebsiella/aislamiento & purificación , Antibacterianos/farmacología , Farmacorresistencia Bacteriana Múltiple/efectos de los fármacos , Escherichia coli/efectos de los fármacos , Humanos , Klebsiella/efectos de los fármacos , Pruebas de Sensibilidad Microbiana , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción
6.
Clin Chem Lab Med ; 56(2): 220-228, 2018 01 26.
Artículo en Inglés | MEDLINE | ID: mdl-28841569

RESUMEN

BACKGROUND: Circulating tumour DNA (ctDNA) is considered to have a high potential for future management of malignancies. This pilot external quality assessment (EQA) scheme aimed to address issues of analytical quality in this new area of laboratory diagnostics. METHODS: The EQA scheme consisted of three 2-mL EDTA-plasma samples spiked with fragmented genomic DNA with a mutant allele frequency ranging from 0% to 10% dedicated to the analysis of nine known sequence variations in KRAS codon 12/13 and of BRAF V600E. Laboratories reported: (1) time elapsed for processing, (2) storage temperatures, (3) methods for extraction and quantification, (4) genotyping methodologies and (5) results. RESULTS: Specimens were sent to 42 laboratories from 10 European countries; 72.3% reported to isolate cell-free DNA (cfDNA) manually, 62.5% used the entire plasma volume for cfDNA isolation and 38.5% used >10% of cfDNA extracted for downstream genotyping. Of the methods used for quantification, PicoGreen demonstrated the lowest coefficient of variation (33.7%). For genotyping, 11 different methods were reported with the highest error rate observed for Sanger sequencing and the lowest for highly sensitive approaches like digital PCR. In total, 197 genotypes were determined with an overall error rate of 6.09%. CONCLUSIONS: This pilot EQA scheme illustrates the current variability in multiple phases of cfDNA processing and analysis of ctDNA resulting in an overall error rate of 6.09%. The areas with the greatest variance and clinical impact included specimen volume, cfDNA quantification method, and preference of genotyping platform. Regarding quality assurance, there is an urgent need for harmonisation of procedures and workflows.


Asunto(s)
Técnicas de Química Analítica/normas , ADN Tumoral Circulante/análisis , ADN Tumoral Circulante/aislamiento & purificación , Técnicas de Genotipaje/normas , Técnicas de Química Analítica/métodos , Técnicas de Genotipaje/métodos , Humanos , Biopsia Líquida , Volumen Plasmático , Manejo de Especímenes , Flujo de Trabajo
7.
Clin Chem Lab Med ; 57(1): 78-88, 2018 12 19.
Artículo en Inglés | MEDLINE | ID: mdl-29729139

RESUMEN

BACKGROUND: The International Organization for Standardization (ISO) 15189 standard provides recommendations for the postexamination reporting phase to enhance quality in clinical laboratories. The purpose of this study was to encourage a broad discussion on current reporting practices for molecular diagnostic tests by conducting a global survey of such practices. METHODS: The International Federation of Clinical Chemistry and Laboratory Medicine's Committee for Molecular Diagnostics (IFCC C-MD) surveyed laboratories on selected ISO 15189 recommendations and topics. The survey addressed the following aspects: (1) laboratory demographics, (2) report format, (3) result reporting/layout, (4) comments in report and (5) interpretation and clinical decision-making information. Additionally, participants indicated categories needing standardization. RESULTS: Sixteen responses from laboratories located in Asia, Europe, the Middle East, North America and South America were received. Several categories yielded 100% agreement between laboratories, whereas other categories had less than or equal to 50% concordance. Participants scored "nomenclature" and "description of methodologies" as the two most frequently cited aspects needing standardization. CONCLUSIONS: The postexamination phase requires extensive and consistent communication between the laboratory, the healthcare provider and the end user. Surveyed laboratories were most likely to follow explicit ISO 15189 recommendations vs. recommendations when the term(s) "where appropriate or where applicable" was used. Interpretation and reporting of critical values varied among participants. Although the outcome of this study may not fully represent the practices of all molecular testing laboratories in countries around the world, the survey identified and specified several recommendations that are requirements for harmonized reporting in molecular diagnostics.


Asunto(s)
Internacionalidad , Técnicas de Diagnóstico Molecular/normas , Encuestas y Cuestionarios , Humanos , Estándares de Referencia
8.
Clin Chem ; 62(8): 1084-95, 2016 08.
Artículo en Inglés | MEDLINE | ID: mdl-27324736

RESUMEN

BACKGROUND: Suboptimal laboratory procedures resulting in genotyping errors, misdiagnosis, or incorrect reporting bear greatly on a patient's health management, therapeutic decisions made on their behalf, and ultimate outcome. Participation in external quality assessment (EQA) is a key element of quality assurance in molecular genetic diagnostics. Therefore, the Reference Institute for Bioanalytics has tried for 13 years to improve the quality of genetic testing by offering an EQA for different clinically relevant sequence variations. METHODS: Within each of the biannual EQA schemes offered, up to 18 samples of lyophilized human genomic DNA were provided for up to 50 different molecular genetic tests. Laboratories were asked to use their routine procedures for genotyping. At least 2 expert peer assessors reviewed the final returns. Data from 2002 to 2014 were evaluated. RESULTS: In total, 82 462 reported results from 812 characterized samples were evaluated. Globally, the number of participants increased each year along with the number of sequence variations offered. The error rate decreased significantly over the years with an overall error rate of 1.44%. Additionally, a decreased error rate for samples repeated over time was noted. Interestingly, the error rate showed a high difference depending on the locus analyzed and the method used. CONCLUSIONS: Based on the evaluation of this long-term EQA scheme, various recommendations can be given to improve the quality of molecular genetic testing, such as the use of 2 different methods for genotyping. Furthermore, some methods are inappropriate for analysis of certain sequence variations.


Asunto(s)
ADN/genética , Pruebas Genéticas/normas , Genotipo , Humanos , Garantía de la Calidad de Atención de Salud
9.
Clin Chem Lab Med ; 53(12): 1927-34, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26053008

RESUMEN

BACKGROUND: Biobanks are becoming increasingly important for assessment of disease risk as well as identification and validation of new diagnostic biomarkers and druggable targets. The validity of data obtained from biobanks is critically limited by the biomaterial quality of the biological samples. External quality assessment (EQA) programs suitable to comprehensively measure the biomaterial quality in archived materials are currently lacking. We report on quantitative assay designs for the analysis of both structural and functional integrity of DNAs that were applied in a first pilot EQA within the priority program on tumor tissue biobanking funded by the German Cancer Aid. METHODS: Participating biobanks isolated DNAs from a standardized set of 10 samples comprising sections of four different formalin-fixed paraffin-embedded tissues using their standard operating procedures. Isolated DNAs and analytical results were returned and analyzed centrally for nucleic acids yield, purity, fragmentation and amplificability at a quantitative level using dedicated assay designs. RESULTS: The amount of extracted DNA varied in isolates ranging between 1.5 µg and 25.8 µg. Quantification of DNA fragmentation and amplificability allowed to highlight considerable discrepancies in DNA quality. Amplicons yielded from the isolates of these identical EQA samples ranged from 105 to 411 bp suggesting differences between residual inhibitors of downstream enzymatic reactions. CONCLUSIONS: The quality of extraction of bioanalytes from biomaterial archives is heterogeneous even for stable biomolecules like DNA isolated with highly standardized methods. EQAs are appropriate tools to uncover strengths and weaknesses in biobanks in a systematic fashion. Biomaterial integrity is insufficiently reflected by standard methods, but needs to be assessed to improve biobank interoperability. Finally, our results also point towards the problem of measuring the quality of more delicate biomolecules like proteins or metabolites.


Asunto(s)
ADN/aislamiento & purificación , Formaldehído/química , Adhesión en Parafina , Bancos de Tejidos/normas , ADN/genética , ADN/normas , Humanos , Adhesión en Parafina/normas , Control de Calidad
10.
BMC Cardiovasc Disord ; 15: 50, 2015 Jun 14.
Artículo en Inglés | MEDLINE | ID: mdl-26072112

RESUMEN

BACKGROUND: To evaluate diagnostic and long-term prognostic values of hFABP compared to NT-proBNP and troponin I (TnI) in patients presenting to the emergency department (ED) suspected of acute heart failure (AHF). METHODS: 401 patients with acute dyspnea or peripheral edema, 122 suffering from AHF, were prospectively enrolled and followed up to 5 years. hFABP combined with NT-proBNP versus NT-proBNP alone was tested for AHF diagnosis. Prognostic value of hFABP versus TnI was evaluated in models predicting all-cause mortality (ACM) and AHF related rehospitalization (AHF-RH) at 1 and 5 years, including 11 conventional risk factors plus NT-proBNP. RESULTS: Additional hFABP measurements improved diagnostic specificity and positive predictive value (PPV) of sole NT-proBNP testing at the cutoff <300 ng/l to "rule out" AHF. Highest hFABP levels (4th quartile) were associated with increased ACM (hazard ratios (HR): 2.1-2.5; p = 0.04) and AHF-RH risk at 5 years (HR 2.8-8.3, p = 0.001). ACM was better characterized in prognostic models including TnI, whereas AHF-RH was better characterized in prognostic models including hFABP. Cox analyses revealed a 2 % increase of ACM risk and 3-7 % increase of AHF-RH risk at 5 years by each unit increase of hFABP of 10 ng/ml. CONCLUSIONS: Combining hFABP plus NT-proBNP (<300 ng/l) only improves diagnostic specificity and PPV to rule out AHF. hFABP may improve prognosis for long-term AHF-RH, whereas TnI may improve prognosis for ACM. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT00143793 .


Asunto(s)
Proteínas de Unión a Ácidos Grasos/sangre , Insuficiencia Cardíaca/sangre , Isquemia Miocárdica/sangre , Péptido Natriurético Encefálico/sangre , Fragmentos de Péptidos/sangre , Troponina I/sangre , Enfermedad Aguda , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Área Bajo la Curva , Estudios de Cohortes , Diagnóstico Diferencial , Disnea/sangre , Disnea/diagnóstico , Disnea/etiología , Ecocardiografía , Edema/sangre , Edema/diagnóstico , Edema/etiología , Servicio de Urgencia en Hospital , Proteína 3 de Unión a Ácidos Grasos , Insuficiencia Cardíaca/complicaciones , Insuficiencia Cardíaca/diagnóstico , Humanos , Estimación de Kaplan-Meier , Estudios Longitudinales , Persona de Mediana Edad , Isquemia Miocárdica/diagnóstico , Readmisión del Paciente , Pronóstico , Estudios Prospectivos , Sensibilidad y Especificidad , Tasa de Supervivencia , Adulto Joven
11.
Clin Lab ; 61(11): 1737-47, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-26732000

RESUMEN

BACKGROUND: To evaluate the diagnostic and prognostic value of sensitive troponin I (TnI) in patients with acute dyspnea and/or peripheral edema suspected of having acute heart failure (AHF). METHODS: This single centre prospective clinical study evaluates 372 patients presenting with acute dyspnea and/or peripheral edema to the emergency department (ED). Measurements of TnI and NT-proBNP were performed at the initial presentation in the ED. All patients were followed up to 5 years. The diagnostic value of TnI compared to NT-proBNP for AHF diagnosis as well as long-term prognostic values for all cause mortality and AHF related rehospitalization were evaluated. RESULTS: TnI plus NT-proBNP improved the diagnosis of AHF (improvement of accuracy (75%, 95% CI 71% - 79%), specificity (68%, 95% CI 62% - 74%), PPV (54%, 95% CI 47% - 62%), and NRI +0.15) compared to NT-proBNP alone (p = 0.0001). TnI levels showed independent prognostic value for all-cause mortality and AHF related rehospitalization after 1 and 5 years (range of AUCs 0.64 - 0.72; p = 0.03 or lower). Highest TnI levels of the 4th quartile revealed an up to 5.5 times higher risk of death within 1 and 5 years (range of HRs: 2.5 - 5.5; p = 0.0001). TnI added significantly to multivariable Cox prediction models even after adjusting for NT-proBNP, particularly in AHF patients (range of HRs: 2.1 - 2.7; p ≤ 0.05). CONCLUSIONS: TnI improves AHF diagnosis when combined with NT-proBNP. TnI identifies patients with high 1- and 5-year all-cause mortality and AHF-related rehospitalization risk and adds prognostic value to NT-proBNP.


Asunto(s)
Insuficiencia Cardíaca/diagnóstico , Troponina I/sangre , Enfermedad Aguda , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Insuficiencia Cardíaca/sangre , Insuficiencia Cardíaca/fisiopatología , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Sensibilidad y Especificidad , Adulto Joven
13.
J Breath Res ; 18(1)2023 12 27.
Artículo en Inglés | MEDLINE | ID: mdl-38100823

RESUMEN

Exhaled breath analysis is evolving into an increasingly important non-invasive diagnostic tool. Volatile organic compounds (VOCs) in breath contain information about health status and are promising biomarkers for several diseases, including respiratory infections caused by bacteria. To monitor the composition of VOCs in breath or the emission of VOCs from bacteria, sensitive analytical techniques are required. Next to mass spectrometry, ion mobility spectrometry (IMS) is considered a promising analytical tool for detecting gaseous analytes in the parts per billion by volume to parts per trillion by volume range. This work presents a new, dual coupling of thermal desorption gas chromatography to a quadrupole mass spectrometer (MS) and an IMS by operating a simple splitter. Nearly identical retention times can be reached in the range of up to 30 min with slight deviations of 0.06 min-0.24 min. This enables the identification of unknown compounds in the IMS chromatogram using unambiguous mass spectral identification, as there are still no commercially available databases for IMS. It is also possible to discriminate one of the detectors using the splitter to improve detection limits. Using a test liquid mixture of seven ketones, namely 2-butanone, 2-pentanone, 2-hexanone, 2-heptanone, 2-octanone, 2-nonanone, and 2-decanone with a concentration of 0.01 g l-1reproducibilities ranging from 3.0% to 7.6% for MS and 2.2%-5.3%, for IMS were obtained, respectively. In order to test the system optimized here for the field of breath analysis, characteristic VOCs such as ethanol, isoprene, acetone, 2-propanol, and 1-propanol were successfully identified in exhaled air using the dual detector system due to the match of the corresponding IMS, and MS spectra. The presented results may be considered to be a starting point for the greater use of IMS in combination with MS within the medical field.


Asunto(s)
Espectrometría de Movilidad Iónica , Compuestos Orgánicos Volátiles , Humanos , Cromatografía de Gases y Espectrometría de Masas/métodos , Pruebas Respiratorias/métodos , Espectrometría de Masas/métodos , Acetona/análisis , Compuestos Orgánicos Volátiles/análisis , Bacterias
14.
Clin Chim Acta ; 547: 117398, 2023 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-37217114

RESUMEN

BACKGROUND: The clinical validity of ctDNA analysis as a diagnostic, prognostic and predictive biomarker has been demonstrated in many studies. The rapid spread of tests for the analysis of ctDNA raises questions regarding their standardization and quality assurance. The aim of this study was to provide a global overview of the test methods, laboratory procedures and quality assessment practices using ctDNA diagnostics. METHODS: The Molecular Diagnostics Committee of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC C-MD) conducted a survey among international laboratories performing ctDNA analysis. Questions on analytical techniques, test parameters, quality assurance and the reporting of findings were included. RESULTS: A total of 58 laboratories participated in the survey. The majority of the participating laboratories (87.7 %) performed testing for patient care. Most laboratories conducted their assays for lung cancer (71.9 %), followed by colorectal (52.6 %) and breast (40.4 %) cancer, and 55.4 % of the labs used ctDNA analysis for follow-up/monitoring of treatment-resistant alterations. The most frequent gene analysed was EGFR (75.8 %), followed by KRAS (65.5 %) and BRAF (56.9 %). Participation in external quality assessment programs was reported by only 45.6 % of laboratories. CONCLUSIONS: The survey indicates that molecular diagnostic methods for the analysis of ctDNA are not standardized across countries and laboratories. Furthermore, it reveals a number of differences regarding sample preparation, processing and reporting test results. Our findings indicate that ctDNA testing is being conducted without sufficient attention to analytical performance between laboratories and highlights the need for standarisation of ctDNA analysis and reporting in patient care.


Asunto(s)
ADN Tumoral Circulante , Neoplasias Pulmonares , Humanos , Laboratorios , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Estándares de Referencia , Pronóstico , Mutación , Biomarcadores de Tumor/genética
15.
Viruses ; 14(3)2022 02 27.
Artículo en Inglés | MEDLINE | ID: mdl-35336898

RESUMEN

Respiratory viruses play an important role in asthma exacerbation, and early exposure can be involved in recurrent bronchitis and the development of asthma. The exact mechanism is not fully clarified, and pathogen-to-host interaction studies are warranted to identify biomarkers of exacerbation in the early phase. Only a limited number of international exacerbation cohorts were studied. Here, we have established a local pediatric exacerbation study in Germany consisting of children with asthma or chronic, recurrent bronchitis and analyzed the viriome within the nasopharyngeal swab specimens derived from the entire cohort (n = 141). Interestingly, 41% of exacerbated children had a positive test result for human rhinovirus (HRV)/human enterovirus (HEV), and 14% were positive for respiratory syncytial virus (RSV). HRV was particularly prevalent in asthmatics (56%), wheezers (50%), and atopic (66%) patients. Lymphocytes were decreased in asthmatics and in HRV-infected subjects, and patients allergic to house dust mites were more susceptible to HRV infection. Our study thus confirms HRV infection as a strong 'biomarker' of exacerbated asthma. Further longitudinal studies will show the clinical progress of those children with a history of an RSV or HRV infection. Vaccination strategies and novel treatment guidelines against HRV are urgently needed to protect those high-risk children from a serious course of disease.


Asunto(s)
Asma , Bronquitis , Infecciones por Enterovirus , Enterovirus , Infecciones por Picornaviridae , Infecciones por Virus Sincitial Respiratorio , Virus Sincitial Respiratorio Humano , Infecciones del Sistema Respiratorio , Virosis , Virus , Asma/epidemiología , Biomarcadores , Niño , Humanos , Lactante , Infecciones del Sistema Respiratorio/epidemiología , Rhinovirus
16.
Clin Chim Acta ; 531: 237-242, 2022 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-35413298

RESUMEN

BACKGROUND: In the current COVID-19 pandemic, early and rapid diagnosis of potentially infected and contagious individuals enables containment of the disease through quarantine and contact tracing. The rapid global expansion of these diagnostic testing services raises questions concerning the current state of the art with regard to standardization of testing and quality assessment practices. The aim of this study was to provide a global overview of the test methods, laboratory procedures and quality assessment practices used for SARS-CoV-2 diagnostics. METHODS: The Molecular Diagnostics Committee of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC C-MD) initiated a survey among international laboratories performing molecular genetic detection of SARS-CoV-2. Questions on quality assurance, variant testing, sequencing and the transmission of findings were included in the survey. RESULTS: A total of 273 laboratories from 49 countries participated in the survey. The majority of the participating laboratories (92.2%) use reverse transcriptase polymerase chain reaction (RT-PCR). The majority of participating laboratories do not conduct testing to identify SARS CoV-2 variants. Participation in external quality assessment programs was reported by the majority of laboratories, however, 33.2% of the laboratories reported not participating in external quality assurance programmes. CONCLUSIONS: Based on the survey, molecular diagnostic methods for SARS-CoV-2 detection are clearly not standardized across different countries and laboratories. The survey found an array of responses in regard to sample preparation, collection, processing and reporting of results. This work suggests quality assurance is insufficiently performed by diagnostic laboratories conducting SARS-CoV-2 testing.


Asunto(s)
COVID-19 , SARS-CoV-2 , COVID-19/diagnóstico , Prueba de COVID-19 , Humanos , Pandemias , Patología Molecular , SARS-CoV-2/genética
17.
J Proteome Res ; 10(10): 4661-70, 2011 Oct 07.
Artículo en Inglés | MEDLINE | ID: mdl-21827211

RESUMEN

The Interleukin-1/Toll-like receptor signaling pathway is a crucial signaling pathway within the innate immune system and the use of mass spectrometric techniques became valuable to investigate signal transduction pathways. To date only a few reports exist that focus on the mass spectrometric identification of novel signaling intermediates within the TLR signal transduction pathway. Here we used this approach systematically to identify new interaction partners of the TLR signaling pathway and subsequently characterized them functionally. We identified 14-3-3 theta as a new member of the TLR signaling complex. With genetic complementation assays, we demonstrate that 14-3-3 negatively regulates TLR2-dependent NF-κB activity and amplifies the TLR4-dependent activation of the transcription factor. While 14-3-3 has no effect on TLR-induced apoptosis in innate immune cells, it controls the release of the inflammatory, IRF3-dependent cytokines like RANTES and IP-10 after stimulation with LPS. Most strikingly, 14-3-3 controls the production of proinflammatory cytokines like IL-6, IL-8, and TNFα in a different manner. Our results identify 14-3-3 theta as a new and important regulatory protein in the TLR signaling suppressing the MyD88-dependent pathway.


Asunto(s)
Proteínas 14-3-3/metabolismo , Regulación de la Expresión Génica , Receptor Toll-Like 2/metabolismo , Animales , Apoptosis , Quimiocina CXCL10/metabolismo , Células HEK293 , Humanos , Inflamación , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Espectrometría de Masas/métodos , Ratones , Modelos Biológicos , FN-kappa B/metabolismo , Transducción de Señal , Receptor Toll-Like 4/metabolismo , Transcripción Genética , Factor de Necrosis Tumoral alfa/metabolismo
18.
J Biol Chem ; 285(47): 36486-94, 2010 Nov 19.
Artículo en Inglés | MEDLINE | ID: mdl-20843814

RESUMEN

The innate immune system employs Toll-like receptors (TLRs) for the detection of invading microorganisms based on distinct molecular patterns. For example, TLR9 is activated by microbial DNA and also by short therapeutic CpG-containing oligonucleotides (CpG-ODN). TLR9 activation leads to the production of interferons and the priming of humoral adaptive immune responses. Unfortunately, the principles of ligand recognition by TLR9 are poorly understood, and genetic variants of TLR9, which may affect its function, have not been characterized systematically on the molecular level. We therefore sought to functionally characterize reported single nucleotide polymorphisms of TLR9 in the HEK293 model system. We discovered that two variants, P99L and M400I, are associated with altered receptor function regarding NF-κB activation and cytokine induction. Our investigations show that for the most functionally impaired variant, P99L, the ability to respond to physiological and therapeutic TLR9 ligands is severely compromised. However, CpG-ODN binding is normal. CpG-ODN recognition by TLR9 thus appears to involve two separate events, CpG-ODN binding and sensing. Our studies highlight Pro-99 as a residue important for the latter process. In genotyping studies, we confirmed that both M400I (rs41308230) and P99L (rs5743844) are relatively rare variants of TLR9. Our data add rs41308230 and rs5743844 to the list of functionally important TLR variants and warrant further research into their relevance for infectious disease susceptibility or responsiveness to CpG-ODN-based therapies.


Asunto(s)
Mutación/genética , Oligodesoxirribonucleótidos/farmacología , Polimorfismo de Nucleótido Simple/genética , Receptor Toll-Like 9/genética , Western Blotting , Células Cultivadas , Técnica del Anticuerpo Fluorescente , Genotipo , Humanos , Inmunoprecipitación , Riñón/citología , Riñón/metabolismo , Luciferasas/metabolismo , Mutagénesis Sitio-Dirigida , FN-kappa B/genética , FN-kappa B/metabolismo , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Receptor Toll-Like 9/metabolismo
19.
Pathogens ; 10(10)2021 Sep 29.
Artículo en Inglés | MEDLINE | ID: mdl-34684208

RESUMEN

In the last two decades, the worldwide dissemination of multidrug-resistant Gram-negative bacteria (MDR-GNB) has continued. Therapy options for such infections caused by MDR-GNB remain scarce, and only few new antimicrobial agents have been granted market approval. Cefiderocol has been approved for the treatment of infections associated with aerobic GNB with limited therapy options. This study evaluated the in vitro efficacy of cefiderocol against carbapenem-non-susceptible clinical GNB isolates from Germany. A total of 115 non-duplicate carbapenem-nonsusceptible GNB isolates, 61 (53.05%) of which were Enterobacterales species and 54 (46.95%) were non-fermenters (Acinetobacter baumanii and Pseudomonas aeruginosa), were investigated for their cefiderocol susceptibility. Minimum inhibitory concentrations (MICs) for cefiderocol were determined by disk diffusion, according to EUCAST (European committee for antimicrobial susceptibility testing). Susceptibility rates were based on EUCAST breakpoints. In the absence of a species-specific breakpoint, pharmacokinetic/-dynamic breakpoints were used. The most common pathogen was A. baumannii (33.91%), followed by Klebsiella pneumoniae (31.3%), P. aeruginosa (13.04%) and Escherichia coli (9.57%). Overall, 83.6% (51/61) of the Enterobacterales and 81.48% (44/54) of the non-fermenters were susceptible towards cefiderocol. In total, 20 species of Enterobacterales and non-fermenting GNB were resistant towards cefiderocol, irrespective of the isolation year (2014 to 2021). Moreover, the majority of the resistant isolates were among the OXA-23 producing A. baumannii (n = 7/26; 26.92%) from patients hospitalized during 2018 and 2019. Cefiderocol demonstrated high in vitro susceptibility rates against a wide range of carbapenem-non-susceptible GNB, including carbapenemase-producing isolates. Cefiderocol exhibited stability against hydrolysis by all carbapenemases, including metallo-ß-lactamases (MBLs), except that few OXA-producing isolates exhibited resistance towards cefiderocol.

20.
Children (Basel) ; 8(8)2021 Aug 08.
Artículo en Inglés | MEDLINE | ID: mdl-34438573

RESUMEN

We assessed the performance of a rapid antigen test (RAT) in everyday clinical practice. Between 1 November 2020 until 1 April 2021 all in-patients at the Helios University Hospital Wuppertal, Germany, as well as the accompanying relatives at the Children's Hospital received a SARS-CoV-2 RAT and a SARS-CoV-2 RT-PCR prior to admission. Out of 3686 patients, 22 (0.6%) subjects were tested positive by RT-PCR and RAT, and 3591 (97.4%) were negative by both methods, showing discordant results: RT-PCR+/RAT- in 58 (1.6%) and RT-PCR-/RAT+ in 15 patients (0.4%). Overall sensitivity and specificity of RAT was 27.5% (95%CI 18.1-38.6%) and 99.6% (95%CI 99.3-99.8%), respectively. The sensitivity was slightly higher in adults (30.4%, 95%CI 18.8-90.9%) than in pediatric subjects (20.8%, 95%CI 7.1-42.2%). False negative RAT had a statistically higher Ct-value (p < 0.001) compared to true positive values, and overall sensitivity increased to 80% [59.3-93.2%] with Ct value < 30. While the sensitivity of the RAT was poor compared with the RT-PCR, the specificity was excellent. However, the sensitivity increased with lower Ct value, and with the right anamnesis the RAT can be a quick and easy approach to distinguish people who are infectious with SARS-CoV-2 from noninfectious people, enabling appropriate triage in clinical practice while waiting for the RT-PCR result.

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