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1.
Ann Med ; 53(1): 34-42, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-32808808

RESUMO

BACKGROUND: Studies have demonstrated the diagnostic efficiency of antibody testing in COVID-19 infection. There is limited data on the IgM/IgG changes in asymptomatic and discharged patients with reoccurring positive nucleic acid test (RPNAT). This study aims to investigate these IgM/IgG changes. METHODS: There were 111 patients with positive nucleic acid test (NAT) and 40 suspected patients enrolled in the study. The serum SARS-CoV-2 specific IgM/IgG antibody levels were retrospectively analysed with the disease progress in asymptomatic and RPNAT patients. RESULTS: The best overall performance was found by combining the IgM, IgG, and CT; 95.1% sensitivity and 75% specificity. This was tested in 111 RT-PCR positive cases. The median IgM and IgG levels were lower in the asymptomatic group compared to the symptomatic group (p < .01). Among 15 RPNAT cases, the IgM levels of the RPNAT group at the time of discharge (IgM2.79, IQR: 0.95-5.37) and retest (IgM 2.35, IQR: 0.88-8.65) were significantly higher than those of the non-reoccurring positive nucleic acid test group (Non-RPNAT) (IgM on discharge: 0.59, IQR: 0.33-1.22, IgG on retest: 0.92, IQR: 0.51-1.58). CONCLUSION: Serum SARS-CoV-2 specific IgM/IgG antibody levels remained at a low level during hospitalisation for asymptomatic patients. Elevated IgM levels may have implications in the identification of RPNAT patients before discharge. Key messages This study determined the IgM/IgG changes in asymptomatic and RPNAT patients. The rate of serum SARS-CoV-2 specific IgM/IgG antibody levels increase in the asymptomatic group was lower than in the symptomatic group during hospitalisation. The IgM level did not decrease significantly at discharge in the RPNAT patients, and was higher than that of the Non-RPNAT group on discharge. These results highlight the importance of timely monitoring of IgM levels to identify RPNAT patients before discharge.


Assuntos
Anticorpos Antivirais/sangue , Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/imunologia , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Pneumonia Viral/imunologia , Estudos de Casos e Controles , Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Feminino , Humanos , Técnicas Imunoenzimáticas/métodos , Masculino , Pandemias , Estudos Retrospectivos
2.
Food Chem ; 337: 127780, 2021 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-32799164

RESUMO

To determine malachite green (MG) and its major metabolite, leucomalachite green (LMG) residual levels in tilapia fish, chemiluminescent enzyme immunoassay (CLEIA) was developed based on a single-chain variable fragment (scFv)-alkaline phosphatase (AP) fusion protein. At first, VH and VL gene sequences were cloned from hybridoma cell lines secreting monoclonal antibody against LMG, and then thoroughly by database-assisted sequence analysis. Finally, the productive VH and VL were assembled to an intact scFv sequence and engineered to produce scFv-AP fusion protein. The fusion protein was further identified as a bifunctional reagent for immunoassay, then a sensitive one-step CLEIA against LMG was developed with a half-maximal inhibitory concentration (IC50) and limit of detection (LOD) of 1.3 and 0.04 ng/mL, respectively. The validation results of this novel competitive CLEIA was in line with those obtained by classical HPLC method for determination of total MG in spiked and field incurred samples.


Assuntos
Produtos Pesqueiros/análise , Técnicas Imunoenzimáticas/métodos , Corantes de Rosanilina/análise , Tilápia , Fosfatase Alcalina/genética , Animais , Anticorpos Monoclonais , Resíduos de Drogas/análise , Hibridomas , Luminescência , Proteínas Recombinantes de Fusão , Anticorpos de Cadeia Única/genética
3.
Zhonghua Yu Fang Yi Xue Za Zhi ; 54(11): 1232-1236, 2020 Nov 06.
Artigo em Chinês | MEDLINE | ID: mdl-33147922

RESUMO

Objective: To evaluate the applicability of limiting antigen avidity enzyme immunoassay (LAg-Avidity EIA, LAg for short) in determining the new HIV-1 infection status of MSM population with seroconversion and make correlation analysis of other biological indicators. Methods: The 15 cases with HIV seroconversion were found in a MSM observation cohort for calculating the HIV prevalence in Zhejiang.The subjects were conducted epidemiological investigation and sampled.The interval of infection time was estimated according to the exposure history and the time of HIV-positive confirmation.LAg, immunoblotting, CD4 cell counting and viral load test were applied in the testing of the related blood samples. McNermar test was conducted for consistency of the two methods. Results: Of 15 cases, the average age was (31.5±8.0) years old, ranging from 24 to 57 years old. The interval of infection time ranged from 40 days to 366 days, and the median was 134 days, with inter-quartile range from 89 to 180 days. A total of 7 cases were classified as new HIV-1 infection by LAg, and 8 cases were classified as chronic infection.The consistent rate was high to 86.67%, and kappa value was 0.73.The samples lacking at least two bands in p31, p51, p66 and gp120 by immunoblotting were determined as recent infection, of which the new infection proportion was significantly higher than that of other samples (P=0.029).There was no statistical difference in the distribution of CD4 counts (P=0.533) and viral loads (P=0.467) between the new infection and chronic infection groups that divided by LAg. Conclusion: By combining with exposure history, the limiting antigen avidity enzyme immunoassay can be used to estimate the new HIV-1 infection.The other biological indicators such as immunoblotting bands, CD4 cell counts and viral loads, can be used as accessory indicators in evaluating the status of new HIV-1 infection.


Assuntos
Infecções por HIV , Soropositividade para HIV , HIV-1 , Técnicas Imunoenzimáticas , Adulto , Anticorpos Anti-HIV , Homossexualidade Masculina , Humanos , Masculino , Pessoa de Meia-Idade , Soroconversão , Carga Viral , Adulto Jovem
4.
PLoS One ; 15(10): e0239782, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33091019

RESUMO

The Mozambique Indicators of Immunization, Malaria and HIV/AIDS (IMASIDA) survey was conducted in 2015 and used a two Enzyme Immunoassay (EIA) (Vironostika HIV-1/2 and Murex HIV-1/2) based algorithm to determine the HIV status of the consented participants. The Mozambique Ministry of Health, with support from the US Centers for Disease Control and Prevention (US CDC), added Bio-Rad Geenius™ HIV-1/2 Supplemental Assay to the IMASIDA HIV testing algorithm to confirm all specimens that were found to be reactive on one or both EIAs. In total 11690 specimens were collected to estimate the proportion of HIV positive samples. Results indicate that the proportion of HIV positive samples based on the concordant positive results of two EIA assays was 21.5% (2518/11690). The addition of the Geenius assay to the IMASIDA HIV testing algorithm demonstrated that 792 (31.5%) of 2518 specimens were false-positive and reduced the proportion of HIV positive samples to 14.7% (1722/11690), demonstrating the importance of including a highly specific HIV test to confirm HIV diagnosis. HIV surveys exclusively based on EIA testing algorithm may result in misleading high prevalence results. Our results demonstrate that more specific confirmatory testing should be added to the EIA-based algorithms to ensure accurate HIV diagnosis and correct HIV prevalence estimate in cross-sectional surveys.


Assuntos
Infecções por HIV/diagnóstico , HIV-1/imunologia , HIV-2/imunologia , Técnicas Imunoenzimáticas , Adolescente , Adulto , Algoritmos , Estudos Transversais , Reações Falso-Positivas , Feminino , Anticorpos Anti-HIV , Infecções por HIV/epidemiologia , Humanos , Lactente , Masculino , Programas de Rastreamento , Pessoa de Meia-Idade , Moçambique , Sensibilidade e Especificidade , Adulto Jovem
5.
Front Immunol ; 11: 570927, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33123144

RESUMO

The emergence and rapid spread of SARS-CoV-2 in December 2019 has brought the world to a standstill. While less pathogenic than the 2002-2003 SARS-CoV, this novel betacoronavirus presents a global threat due to its high transmission rate, ability to invade multiple tissues, and ability to trigger immunological hyperactivation. The identification of the animal reservoir and intermediate host were important steps toward slowing the spread of disease, and its genetic similarity to SARS-CoV has helped to determine pathogenesis and direct treatment strategies. The exponential increase in cases has necessitated fast and reliable testing procedures. Although RT-PCR remains the gold standard, it is a time-consuming procedure, paving the way for newer techniques such as serologic tests and enzyme immunoassays. Various clinical trials using broad antiviral agents in addition to novel medications have produced controversial results; however, the advancement of immunotherapy, particularly monoclonal antibodies and immune modulators is showing great promise in clinical trials. Non-orthodox medications such as anti-malarials have been tested in multiple institutions but definitive conclusions are yet to be made. Adjuvant therapies have also proven to be effective in decreasing mortality in the disease course. While no formal guidelines have been established, the multitude of ongoing clinical trials as a result of unprecedented access to research data brings us closer to halting the SARS-CoV-2 pandemic.


Assuntos
Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus , Pandemias , Pneumonia Viral , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/uso terapêutico , Alanina/análogos & derivados , Alanina/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Anticorpos Antivirais/uso terapêutico , Betacoronavirus/genética , Betacoronavirus/imunologia , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/patologia , Infecções por Coronavirus/terapia , Reservatórios de Doenças/virologia , Reposicionamento de Medicamentos/métodos , Humanos , Técnicas Imunoenzimáticas , Imunoterapia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/diagnóstico , Pneumonia Viral/patologia , Pneumonia Viral/terapia , Receptores Virais/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Testes Sorológicos/métodos , Glicoproteína da Espícula de Coronavírus/metabolismo
6.
BMC Infect Dis ; 20(1): 740, 2020 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-33036575

RESUMO

BACKGROUND: From 2016, the Government of India introduced the oral rotavirus vaccine into the national immunization schedule. Currently, two indigenously developed vaccines (ROTAVAC, Bharat Biotech; ROTASIIL, Serum Institute of India) are included in the Indian immunization program. We report the rotavirus disease burden and the diversity of rotavirus genotypes from 2005 to 2016 in a multi-centric surveillance study before the introduction of vaccines. METHODS: A total of 29,561 stool samples collected from 2005 to 2016 (7 sites during 2005-2009, 3 sites from 2009 to 2012, and 28 sites during 2012-2016) were included in the analysis. Stools were tested for rotavirus antigen using enzyme immunoassay (EIA). Genotyping was performed on 65.8% of the EIA positive samples using reverse transcription- polymerase chain reaction (RT-PCR) to identify the G (VP7) and P (VP4) types. Multinomial logistic regression was used to quantify the odds of detecting genotypes across the surveillance period and in particular age groups. RESULTS: Of the 29,561 samples tested, 10,959 (37.1%) were positive for rotavirus. There was a peak in rotavirus positivity during December to February across all sites. Of the 7215 genotyped samples, G1P[8] (38.7%) was the most common, followed by G2P[4] (12.3%), G9P[4] (5.8%), G12P[6] (4.2%), G9P[8] (4%), and G12P[8] (2.4%). Globally, G9P[4] and G12P[6] are less common genotypes, although these genotypes have been reported from India and few other countries. There was a variation in the geographic and temporal distribution of genotypes, and the emergence or re-emergence of new genotypes such as G3P[8] was seen. Over the surveillance period, there was a decline in the proportion of G2P[4], and an increase in the proportion of G9P[4]. A higher proportion of mixed and partially typed/untyped samples was also seen more in the age group 0-11 months. CONCLUSIONS: This 11 years surveillance highlights the high burden of severe rotavirus gastroenteritis in Indian children < 5 years of age before inclusion of rotavirus vaccines in the national programme. Regional variations in rotavirus epidemiology were seen, including the emergence of G3P[8] in the latter part of the surveillance. Having pre-introduction data is important to track changing epidemiology of rotaviruses, particularly following vaccine introduction.


Assuntos
Gastroenterite/epidemiologia , Genótipo , Hospitalização , Infecções por Rotavirus/epidemiologia , Rotavirus/genética , Doença Aguda , Antígenos Virais/imunologia , Pré-Escolar , Fezes/virologia , Feminino , Gastroenterite/prevenção & controle , Gastroenterite/virologia , Técnicas de Genotipagem , Humanos , Programas de Imunização , Esquemas de Imunização , Técnicas Imunoenzimáticas , Índia/epidemiologia , Lactente , Recém-Nascido , Masculino , Prevalência , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Rotavirus/imunologia , Infecções por Rotavirus/prevenção & controle , Infecções por Rotavirus/virologia , Vacinas contra Rotavirus/imunologia
8.
Int J Infect Dis ; 99: 397-402, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32800855

RESUMO

In routine clinical practice, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is determined by reverse-transcription PCR (RT-PCR). In the current pandemic, a more rapid and high-throughput method is in growing demand. Here, we validated the performance of a new antigen test (LUMIPULSE) based on chemiluminescence enzyme immunoassay. A total of 313 nasopharyngeal swabs (82 serial samples from 7 infected patients and 231 individual samples from 4 infected patients and 215 uninfected individuals) were analyzed for SARS-CoV-2 with quantitative RT-PCR (RT-qPCR) and then subjected to LUMIPULSE. We determined the cutoff value for antigen detection using receiver operating characteristic curve analysis and compared the performance of the antigen test with that of RT-qPCR. We also compared the viral loads and antigen levels in serial samples from seven infected patients. Using RT-qPCR as the reference, the antigen test exhibited 55.2% sensitivity and 99.6% specificity, with a 91.4% overall agreement rate (286/313). In specimens with > 100 viral copies and between 10 and 100 copies, the antigen test showed 100% and 85% concordance with RT-qPCR, respectively. This concordance declined with lower viral loads. In the serially followed patients, the antigen levels showed a steady decline, along with viral clearance. This gradual decline was in contrast with the abrupt positive-to-negative and negative-to-positive status changes observed with RT-qPCR, particularly in the late phase of infection. In summary, the LUMIPULSE antigen test can rapidly identify SARS-CoV-2-infected individuals with moderate to high viral loads and may be helpful for monitoring viral clearance in hospitalized patients.


Assuntos
Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Reação em Cadeia da Polimerase em Tempo Real , Betacoronavirus , Técnicas de Laboratório Clínico/métodos , Humanos , Técnicas Imunoenzimáticas , Medições Luminescentes , Cavidade Nasal/virologia , Pandemias , Reação em Cadeia da Polimerase em Tempo Real/métodos , Sensibilidade e Especificidade , Carga Viral
9.
Artigo em Inglês | MEDLINE | ID: mdl-32609256

RESUMO

Coronavirus disease 2019 (COVID-19) is an infectious disease initially reported in China and currently worldwide dispersed caused by a new coronavirus (SARS-CoV-2 or 2019-nCoV) affecting more than seven million people around the world causing more than 400 thousand deaths (on June 8th, 2020). The diagnosis of COVID-19 is based on the clinical and epidemiological history of the patient. However, the gold standard for COVID-19 diagnosis is the viral detection through the amplification of nucleic acids. Although the quantitative Reverse-Transcription Polymerase Chain Reaction (RT-PCR) has been described as the gold standard for diagnosing COVID-19, there are several difficulties involving its use. Here we comment on RT-PCR and describe alternative tests developed for the diagnosis of COVID-19.


Assuntos
Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/epidemiologia , Pandemias , Pneumonia Viral/diagnóstico , Pneumonia Viral/epidemiologia , Betacoronavirus/genética , Ensaio de Imunoadsorção Enzimática/métodos , Humanos , Imunoensaio/métodos , Técnicas Imunoenzimáticas/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , Técnicas de Amplificação de Ácido Nucleico/normas , Análise Serial de Proteínas/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Avaliação de Sintomas/métodos
10.
J Clin Microbiol ; 58(10)2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32665420

RESUMO

Coronavirus disease (COVID) serological tests are essential to determine the overall seroprevalence of a population and to facilitate exposure estimates within that population. We performed a head-to-head assessment of enzyme immunoassays (EIAs) and point-of-care lateral flow assays (POCTs) to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies. Demographics, symptoms, comorbidities, treatment, and mortality of patients whose sera were used were also reviewed. Six EIAs (Abbott, Affinity, Bio-Rad, DiaSorin, Euroimmun, and Roche) and six POCTs (BTNX, Biolidics, Deep Blue, Genrui, Getein BioTech, and Innovita) were evaluated for the detection of SARS-CoV-2 antibodies in known COVID-19-infected individuals. Sensitivity of EIAs ranged from 50 to 100%, with only four assays having overall sensitivities of >95% after 21 days after symptom onset. Notably, cross-reactivity with other respiratory viruses (parainfluenza virus [PIV-4] [n = 5], human metapneumovirus [hMPV] [n = 3], rhinovirus/enterovirus [n = 1], CoV-229E [n = 2], CoV-NL63 [n = 2], and CoV-OC43 [n = 2]) was observed; however, overall specificity of EIAs was good (92 to 100%; all but one assay had specificity above 95%). POCTs were 0 to 100% sensitive >21 days after onset, with specificity ranging from 96 to 100%. However, many POCTs had faint banding and were often difficult to interpret. Serology assays can detect SARS-CoV-2 antibodies as early as 10 days after symptom onset. Serology assays vary in their sensitivity based on the marker (IgA/IgM versus IgG versus total) and by manufacturer; however, overall only 4 EIAs and 4 POCTs had sensitivities of >95% >21 days after symptom onset. Cross-reactivity with other seasonal coronaviruses is of concern. Serology assays should not be used for the diagnosis of acute infection but rather in carefully designed serosurveys to facilitate understanding of seroprevalence in a population and to identify previous exposure to SARS-CoV-2.


Assuntos
Anticorpos Antivirais/sangue , Betacoronavirus/isolamento & purificação , Adulto , Idoso , Idoso de 80 Anos ou mais , Betacoronavirus/imunologia , Técnicas de Laboratório Clínico , Infecções por Coronavirus/sangue , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/epidemiologia , Reações Cruzadas , Feminino , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Sistemas Automatizados de Assistência Junto ao Leito , Sensibilidade e Especificidade , Estudos Soroepidemiológicos , Testes Sorológicos , Fatores de Tempo
11.
Viruses ; 12(6)2020 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-32466458

RESUMO

The recent outbreak of the Coronavirus disease 2019 (COVID-19) has quickly spread worldwide since its discovery in Wuhan city, China in December 2019. A comprehensive strategy, including surveillance, diagnostics, research, clinical treatment, and development of vaccines, is urgently needed to win the battle against COVID-19. The past three unprecedented outbreaks of emerging human coronavirus infections at the beginning of the 21st century have highlighted the importance of readily available, accurate, and rapid diagnostic technologies to contain emerging and re-emerging pandemics. Real-time reverse transcriptase-polymerase chain reaction (rRT-PCR) based assays performed on respiratory specimens remain the gold standard for COVID-19 diagnostics. However, point-of-care technologies and serologic immunoassays are rapidly emerging with high sensitivity and specificity as well. Even though excellent techniques are available for the diagnosis of symptomatic patients with COVID-19 in well-equipped laboratories; critical gaps still remain in screening asymptomatic people who are in the incubation phase of the virus, as well as in the accurate determination of live viral shedding during convalescence to inform decisions for ending isolation. This review article aims to discuss the currently available laboratory methods and surveillance technologies available for the detection of COVID-19, their performance characteristics and highlight the gaps in current diagnostic capacity, and finally, propose potential solutions. We also summarize the specifications of the majority of the available commercial kits (PCR, EIA, and POC) for laboratory diagnosis of COVID-19.


Assuntos
Técnicas de Laboratório Clínico , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Infecções Assintomáticas , Betacoronavirus , Humanos , Técnicas Imunoenzimáticas , Testes de Neutralização , Técnicas de Amplificação de Ácido Nucleico , Pandemias , Testes Imediatos , Kit de Reagentes para Diagnóstico/normas , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e Especificidade , Testes Sorológicos , Tomografia Computadorizada por Raios X , Eliminação de Partículas Virais
12.
Klin Lab Diagn ; 65(6): 358-361, 2020.
Artigo em Russo | MEDLINE | ID: mdl-32459894

RESUMO

The serum of 1006 children aged 0-18 years and elderly people aged 60 to 90 years and older for the presence of specific class G immunoglobulins to the Epstein-Barr virus was studied using enzyme immunoassay. The dependence of seropositivity of children on their age and seropositivity of more than 98% of all surveyed elderly people is shown.


Assuntos
Infecções por Vírus Epstein-Barr/epidemiologia , Herpesvirus Humano 4 , Adolescente , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/sangue , Criança , Pré-Escolar , Humanos , Técnicas Imunoenzimáticas , Imunoglobulina G/sangue , Lactente , Recém-Nascido , Pessoa de Meia-Idade , Moscou/epidemiologia
13.
J Infect Dis ; 222(2): 189-193, 2020 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-32382737

RESUMO

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel ß-coronavirus, causes severe pneumonia and has spread throughout the globe rapidly. The disease associated with SARS-CoV-2 infection is named coronavirus disease 2019 (COVID-19). To date, real-time reverse-transcription polymerase chain reaction (RT-PCR) is the only test able to confirm this infection. However, the accuracy of RT-PCR depends on several factors; variations in these factors might significantly lower the sensitivity of detection. METHODS: In this study, we developed a peptide-based luminescent immunoassay that detected immunoglobulin (Ig)G and IgM. The assay cutoff value was determined by evaluating the sera from healthy and infected patients for pathogens other than SARS-CoV-2. RESULTS: To evaluate assay performance, we detected IgG and IgM in the sera from confirmed patients. The positive rate of IgG and IgM was 71.4% and 57.2%, respectively. CONCLUSIONS: Therefore, combining our immunoassay with real-time RT-PCR might enhance the diagnostic accuracy of COVID-19.


Assuntos
Anticorpos Antivirais/sangue , Betacoronavirus/imunologia , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Técnicas Imunoenzimáticas/métodos , Pneumonia Viral/diagnóstico , Testes Sorológicos/métodos , Adulto , Infecções por Coronavirus/imunologia , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Medições Luminescentes , Masculino , Pessoa de Meia-Idade , Pandemias , Peptídeos/imunologia , Pneumonia Viral/imunologia , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e Especificidade , Proteínas Virais/imunologia
15.
Euro Surveill ; 25(12)2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32234120

RESUMO

BackgroundTick-borne encephalitis (TBE) is a potentially severe neurological disease caused by TBE virus (TBEV). In Europe and Asia, TBEV infection has become a growing public health concern and requires fast and specific detection.AimIn this observational study, we evaluated a rapid TBE IgM test, ReaScan TBE, for usage in a clinical laboratory setting.MethodsPatient sera found negative or positive for TBEV by serological and/or molecular methods in diagnostic laboratories of five European countries endemic for TBEV (Estonia, Finland, Slovenia, the Netherlands and Sweden) were used to assess the sensitivity and specificity of the test. The patients' diagnoses were based on other commercial or quality assured in-house assays, i.e. each laboratory's conventional routine methods. For specificity analysis, serum samples from patients with infections known to cause problems in serology were employed. These samples tested positive for e.g. Epstein-Barr virus, cytomegalovirus and Anaplasma phagocytophilum, or for flaviviruses other than TBEV, i.e. dengue, Japanese encephalitis, West Nile and Zika viruses. Samples from individuals vaccinated against flaviviruses other than TBEV were also included. Altogether, 172 serum samples from patients with acute TBE and 306 TBE IgM negative samples were analysed.ResultsCompared with each laboratory's conventional methods, the tested assay had similar sensitivity and specificity (99.4% and 97.7%, respectively). Samples containing potentially interfering antibodies did not cause specificity problems.ConclusionRegarding diagnosis of acute TBEV infections, ReaScan TBE offers rapid and convenient complementary IgM detection. If used as a stand-alone, it can provide preliminary results in a laboratory or point of care setting.


Assuntos
Vírus da Encefalite Transmitidos por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/diagnóstico , Imunoglobulina M/sangue , Anticorpos Antivirais/sangue , Encefalite Transmitida por Carrapatos/epidemiologia , Encefalite Transmitida por Carrapatos/imunologia , Feminino , Humanos , Técnicas Imunoenzimáticas , Masculino , Valor Preditivo dos Testes , Sensibilidade e Especificidade
16.
J Steroid Biochem Mol Biol ; 201: 105685, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32320758

RESUMO

During the past 25 years or so a number of studies have been carried out to address the hypothesis that the ratio of 2-hydroxyestrone (2-hydroxy-E1) to 16α-hydroxyestrone (16α-hydroxy-E1) is associated with breast cancer risk. The rationale for this hypothesis is based on data from studies that suggest a tumorigenic and genotoxic effect of 16α-hydroxy-E1 and a protective effect of 2-hydroxy-E1 regarding breast cancer risk. The adverse effect of 16α-hydroxy-E1 has been attributed to its potential to form covalent adducts with macromolecules. Initial studies used radiometric assays and enzyme immunoassays to test the hypothesis. However, concerns about the accuracy of these assays led to the development of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay that is capable of measuring 5 unconjugated and 15 conjugated endogenous estrogens, which include 2- and 16-hydroxylated estrogen metabolites, in serum or urine. The conjugated estrogens are quantified following a deconjugation (hydrolysis) step to remove the sulfate and glucuronide groups. Epidemiologic studies have been using the LC-MS/MS assay to determine whether there is an association between breast cancer risk and the ratio of the sum of the concentrations of metabolites in the 2-hydroxylated estrogen pathway and in the 16-hydroxylated estrogen pathway. However, the validity of the pathways as biomarkers was not evaluated. The 16-hydroxylated estrogen pathway includes estriol, 16-epiestriol, 17-epiestriol and 16-ketoestradiol, in addition to 16α-hydroxy-E1. However, with the exception of 16α-hydroxy-E1, there is no evidence that any of the other estrogens in the pathway have tumorigenic or genotoxic properties, and they do not form covalent adducts with macromolecules. Another deficiency in the epidemiological studies pertains to the accuracy of estrogen metabolite measurements obtained after the hydrolysis step in the LC-MS/MS assays. No validation was performed to demonstrate that a constant efficiency of hydrolysis is found for all the different structural forms of sulfated and glucuronidated conjugates. Other deficiencies in the assays include the need for greater sensitivity so that the very low concentrations of unconjugated 2-hydroxy-E1, 2-hydroxy-E2, and 16α-hydroxy-E1 can be measured in serum. There is also a need to develop assays to measure intact forms of conjugated estrogens in both serum and urine, particularly the sulfates and glucuronides of 2-hydroxylated, 2-methoxylated, and 16α-hydroxylated E1 and E2. This will avoid inaccuracies that stem from hydrolysis procedures. Improvements in LC-MS/MS assay methodology to obtain accurate measurements of unconjugated and conjugated 2-hydroxylated, 2-methoxylated, and 16α-hydroxylated estrogen metabolites are needed. This should provide valuable data for testing the 2-/16α-hydroxylated estrogen-breast cancer risk hypothesis.


Assuntos
Neoplasias da Mama/metabolismo , Hidroxiestronas/metabolismo , Animais , Cromatografia Líquida , Feminino , Humanos , Técnicas Imunoenzimáticas , Ensaio Radioligante , Risco , Espectrometria de Massas em Tandem
17.
Clin Chem Lab Med ; 58(7): 1081-1088, 2020 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-32301749

RESUMO

Background Coronavirus disease 2019, abbreviated to COVID-19, represents an emerging health threat worldwide as, after initial reports in China, it has continued to spread rapidly. The clinical spectrum of the disease varies from mild to severe acute respiratory distress syndrome (ARDS). Moreover, many patients can be asymptomatic, thus increasing the uncertainty of the diagnostic work-up. Laboratory tests play a pivotal role in the diagnosis and management of COVID-19, the current gold standard being real-time reverse transcription polymerase chain reaction (rRT-PCR) on respiratory tract specimens. However, the diagnostic accuracy of rRT-PCR depends on many pre-analytical and analytical variables. The measurement of specific COVID-19 antibodies (both IgG and IgM) should serve as an additional, non-invasive tool for disease detection and management. Methods The imprecision of the MAGLUMI™ 2000 Plus 2019-nCov IgM and IgG assays (Snibe, Shenzhen, China) was assessed by adopting the Clinical and Laboratory Standards Institute (CLSI) EP15-A3 protocol. Linearity of dilution and recovery was evaluated by means of mixes of high-level pools and low-level pools of serum samples. Immunoglobulin time kinetics were evaluated using a series of serum samples, repeatedly collected from COVID-19-positive patients at different times, from <5 days up to 26-30 days. Results Findings at the analytical validation of the assay carried out according to the CLSI EP15-A3 guideline demonstrated that imprecision and repeatability were acceptable (repeatability was <4% and <6% for IgM and IgG, respectively, whilst intermediate imprecision was <6%). In addition, results of dilution and recovery studies were satisfactory. The kinetics of COVID-19 antibodies confirmed previously reported findings, showing a rapid increase of both IgM and IgG after 6-7 days from the symptom onset. IgG had 100% sensitivity on day 12, whilst 88% was the higher positive rate achieved for IgM after the same time interval. Conclusions The findings of this study demonstrate the validity of the MAGLUMI 2000 Plus CLIA assay for the measurement of specific IgM and IgG in sera of COVID-19 patients, and for obtaining valuable data on the kinetics of both (IgM and IgG) COVID-19 antibodies. These data represent a pre-requisite for the appropriate utilization of specific antibodies for the diagnosis and management of COVID-19 patients.


Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Técnicas Imunoenzimáticas/métodos , Pneumonia Viral/imunologia , Anticorpos Antivirais/sangue , China , Técnicas de Laboratório Clínico/métodos , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina M/sangue , Imunoglobulina M/imunologia , Cinética , Medições Luminescentes/métodos , Pandemias , Reação em Cadeia da Polimerase em Tempo Real
20.
Gen Comp Endocrinol ; 293: 113494, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32333913

RESUMO

Integrative behavioral ecology requires accurate and non-invasive measures of hormone mediators for the study of wild animal populations. Biologically sensitive assay systems for the measurement of hormones and their metabolites need to be validated for the species and sample medium (e.g. urine, feces, saliva) of interest. Where more than one assay is available for hormone (metabolite) measurement, antibody selection is useful in identifying the assay that tracks changes in an individuals endocrine activity best, i.e., the most biologically sensitive assay. This is particularly important when measuring how glucocorticoids (GCs) respond to the subtle, additive effects of acute stressors during a predictable metabolic challenge, such as gestation. Here, we validate a group-specific enzyme immunoassay, measuring immunoreactive 11ß-hydroxyetiocholanolone, for use in a wild primate, geladas (Theropithecus gelada). This group-specific assay produced values correlated with those from a previously validated double-antibody, corticosterone 125I radioimmunoassay. However, the results with the group-specific assay showed a stronger response to an ACTH challenge and identified greater variation in gelada immunoreactive fecal glucocorticoid metabolites (iGCMs) compared with the corticosterone assay, indicating a higher biological sensitivity for assessing adrenocortical activity. We then used the group-specific assay to: (1) determine the normative pattern of iGCM levels across gelada gestation, and (2) identify the ecological, social, and individual factors that influence GC output for pregnant females. Using a general additive mixed model, we found that higher iGCM levels were associated with low rank (compared to high rank) and first time mothers (compared to multiparous mothers). This study highlights the importance of assay selection and the efficacy of group-specific assays for hormonal research in non-invasively collected samples. Additionally, in geladas, our results identify some of the factors that increase GC output over and above the already-elevated GC concentrations associated with gestation. In the burgeoning field of maternal stress, these factors can be examined to identify the effects that GC elevations may have on offspring development.


Assuntos
Fezes/química , Glucocorticoides/metabolismo , Metaboloma , Paridade , Theropithecus/metabolismo , Animais , Animais Selvagens/metabolismo , Corticosterona/metabolismo , Feminino , Técnicas Imunoenzimáticas , Masculino , Modelos Biológicos , Gravidez , Radioimunoensaio , Reprodutibilidade dos Testes
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