RESUMO
Serological testing for anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies is used to detect ongoing or past SARS-CoV-2 infections. To study the kinetics of anti-SARS-CoV-2 antibodies and to assess the diagnostic performances of eight serological assays, we used 129 serum samples collected on known days post symptom onset (dpso) from 42 patients with polymerase chain reaction-confirmed coronavirus disease 2019 (COVID-19) and 54 serum samples from healthy blood donors, and children infected with seasonal coronaviruses. The sera were analyzed for the presence of immunoglobulin G (IgG), immunoglobulin M (IgM), and immunoglobulin A (IgA) antibodies using indirect immunofluorescence testing (IIFT) based on SARS-CoV-2-infected cells. They were further tested for antibodies against the S1 domain of the SARS-CoV-2 spike protein (IgG, IgA) and against the viral nucleocapsid protein (IgG, IgM) using enzyme-linked immunosorbent assays. The assay specificities were 94.4%-100%. The sensitivities varied largely between assays, reflecting their respective purposes. The sensitivities of IgA and IgM assays were the highest between 11 and 20 dpso, whereas the sensitivities of IgG assays peaked between 20 and 60 dpso. IIFT showed the highest sensitivities due to the use of the whole SARS-CoV-2 as substrate and provided information on whether or not the individual has been infected with SARS-CoV-2. Enzyme-linked immunosorbent assays provided further information about both the prevalence and concentration of specific antibodies against selected antigens of SARS-CoV-2.
Assuntos
Anticorpos Antivirais/sangue , Teste Sorológico para COVID-19 , COVID-19/sangue , SARS-CoV-2/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/imunologia , COVID-19/diagnóstico , Proteínas do Nucleocapsídeo de Coronavírus/imunologia , Ensaio de Imunoadsorção Enzimática , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Isotipos de Imunoglobulinas/sangue , Cinética , Masculino , Pessoa de Meia-Idade , Fosfoproteínas/imunologia , SARS-CoV-2/isolamento & purificação , Sensibilidade e Especificidade , Glicoproteína da Espícula de Coronavírus/imunologiaRESUMO
OBJECTIVES: During the current pandemic, antibody testing based on venous serum helps to determine whether the tested person has been previously infected with SARS-CoV-2. Alternatively, capillary blood can be taken via a finger prick (dried blood spots, DBS). In this study, paired DBS and venipuncture samples were tested using two serological assays to evaluate the usability of DBS for the detection of anti-SARS-CoV-2 antibodies. METHODS: Paired samples of DBS and venous serum were collected from 389 volunteers, of whom 75 had a recent PCR-confirmed SARS-CoV-2 infection, and tested for anti-SARS-CoV-2 IgG antibodies against both viral S1 and nucleocapsid protein (NCP) antigens using two ELISAs. Degree of agreement and correlation coefficients between ELISA results based on the two sampling methods were calculated. RESULTS: Results of DBS showed almost perfect agreement and high correlations with results from corresponding serum samples in both the S1-based ELISA and the NCP-based ELISA. CONCLUSIONS: ELISA results derived from DBS showed very high agreement to those obtained with serum, supposing adequate usability and robustness of DBS as sample material for detection of anti-SARS-CoV-2 antibodies. In the near future, large-scale epidemiological screening for antibodies against SARS-CoV-2 will be carried out. Since DBS reduce the strain on healthcare institutions regarding sample collection, they have a potential to facilitate efficient community- and population-based screening in the current SARS-CoV-2 pandemic.
Assuntos
Anticorpos Antivirais/sangue , COVID-19/diagnóstico , SARS-CoV-2/imunologia , Anticorpos Antivirais/imunologia , COVID-19/sangue , Teste Sorológico para COVID-19/estatística & dados numéricos , Teste em Amostras de Sangue Seco/estatística & dados numéricos , Ensaio de Imunoadsorção Enzimática/estatística & dados numéricos , Humanos , Modelos Lineares , Manejo de EspécimesRESUMO
BACKGROUND: Maternal acute primary cytomegalovirus (CMV) infection during the first trimester may cause severe long-term sequelae in newborns. For risk assessment, serological screening is routinely performed in pregnant women based on IgM, IgG and avidity tests using whole-virus antigen. A recent study evaluated the diagnostic value of recombinant protein-based ELISAs as second-line tests in pregnancy CMV screening, including anti-p52 IgM and anti-gB IgG as markers defining the early and late phase of infection, respectively. In the present study, these recombinant ELISAs were used as first-line screening tests in daily laboratory routine and compared to lysate-based assays with respect to [i] the number of conclusive results obtained with the initial sample and [ii] the underlying workload. METHODS: 553 unselected routine serum samples from pregnant women were tested for anti-CMV IgM and IgG antibodies using lysate-based ELISAs and avidity testing. Anti-CMV IgM antibodies against recombinant p52 and anti-CMV IgG antibodies against recombinant glycoprotein B (gB) were also determined by ELISA. All assays were performed and interpreted according to the manufacturer's instructions. RESULTS: For lysate-based IgM, IgG and avidity testing, 84.6 % of samples yielded conclusive results in a total of 1156 tests, while 15.4 % needed follow-up testing of a consecutive sample. Anti-p52 CMV IgM and anti-gB CMV IgG testing produced conclusive results for 92.8 % of samples in a total of 1026 tests, while 7.2 % samples required follow-up testing. CONCLUSIONS: The first-line use of ELISAs measuring anti-p52 CMV IgM and anti-gB CMV IgG antibodies to test for maternal CMV infection increases the number of conclusive results derived from an initial serum sample while requiring a considerably lower number of tests compared to the lysate-based approach. For day-to-day routines in a diagnostic laboratory, this high efficiency of the recombinant testing approach has significant practical relevance.
Assuntos
Infecções por Citomegalovirus , Complicações Infecciosas na Gravidez , Gravidez , Feminino , Recém-Nascido , Humanos , Citomegalovirus , Complicações Infecciosas na Gravidez/diagnóstico , Imunoglobulina M , Afinidade de Anticorpos , Infecções por Citomegalovirus/diagnóstico , Anticorpos Antivirais , Testes Sorológicos/métodos , Imunoglobulina G , Proteínas RecombinantesRESUMO
Crimean-Congo Hemorrhagic Fever Virus (CCHFV) is a geographically widespread tick-borne arbovirus that has been recognized by the WHO as an emerging pathogen needing urgent attention to ensure preparedness for potential outbreaks. Therefore, availability of accurate diagnostic tools for identification of acute cases is necessary. A panel comprising 121 sequential serum samples collected during acute, convalescent and subsided phase of PCR-proven CCHFV infection from 16 Kosovar patients was used to assess sensitivity. Serum samples from 60 healthy Kosovar blood donors were used to assess specificity. All samples were tested with two IgM/IgG immunofluorescence assays (IFA) from BNITM, the CCHFV Mosaic 2 IgG and IgM indirect immunofluorescence tests (IIFT) from EUROIMMUN, two BlackBox ELISAs for the detection of CCHFV-specific IgM and IgG antibodies (BNITM), two Anti-CCHFV ELISAs IgM and IgG from EUROIMMUN using recombinant structural proteins of CCHFV antigens, and two ELISAs from Vector-Best (IgM: µ-capture ELISA, IgG: indirect ELISA using immobilized CCHFV antigen). Diagnostic performances were compared between methods using sensitivity, specificity, concordance and degree of agreement with particular focus on the phase of the infection. In early and convalescent phases of infection, the sensitivities for detecting specific IgG antibodies differed for the ELISA test. The BlackBox IgG ELISA yielded the highest, followed by the EUROIMMUN IgG ELISA and finally the VectorBest IgG ELISA with the lowest sensitivities. In the subsided phase, the VectorBest IgM ELISA detected a high rate of samples that were positive for anti-CCHFV IgM antibodies. Both test systems based on immunofluorescence showed an identical sensitivity for detection of anti-CCHFV IgM antibodies in acute and convalescent phases of infection. Available serological test systems detect anti-CCHFV IgM and IgG antibodies accurately, but their diagnostic performances vary with respect to the phase of the infection.
Assuntos
Anticorpos Antivirais/sangue , Ensaio de Imunoadsorção Enzimática/métodos , Técnica Direta de Fluorescência para Anticorpo/métodos , Técnica Indireta de Fluorescência para Anticorpo/métodos , Vírus da Febre Hemorrágica da Crimeia-Congo/imunologia , Febre Hemorrágica da Crimeia/diagnóstico , Adolescente , Adulto , Animais , Criança , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Kosovo , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Doenças Transmitidas por Carrapatos/diagnóstico , Doenças Transmitidas por Carrapatos/virologia , Carrapatos/virologia , Adulto JovemRESUMO
The parasitic roundworm Trichinella spiralis is most commonly transmitted to humans through consumption of raw or undercooked meat of infected pigs or game. To prevent human infection, slaughterhouses perform meat safety surveillance using the gold standard "Magnetic Stirrer Method". We introduce a fast and objective method using automated detection of specific Trichinella spiralis antigens by a newly developed immunoassay based on chemiluminescence (ChLIA). Panel A comprised muscle tissue samples from non-infected pigs (n = 37). Panel B comprised muscle tissue samples from non-infected pigs spiked with different amounts of Trichinella larvae without collagen capsules (n = 56). Panel C contained muscle tissue samples from experimentally infected pigs including Trichinella larvae encapsulated in collagen (n = 32). Each sample was shredded with PBS buffer in a knife mill, destroying Trichinella larvae. Following centrifugation, the supernatant (muscle tissue extract containing released excretory and secretory Trichinella spiralis antigens) was used for Trichinella-specific antigen detection by the new Trichinella ChLIA. The overall accuracy of the Trichinella ChLIA was 97.6 %. The specificity of the Trichinella ChLIA was 100 % (panel A). The sensitivity in samples from experimentally infected pigs was 100 % representing a detection limit of 0.01 larvae per gram. Cross-reactivity with parasites other than Trichinella spp. was not observed. This new meat inspection method for the detection of Trichinella spiralis antigens presents high specificity and high sensitivity, especially in truly infected samples. In contrast to the gold standard, this new approach to meat safety surveillance does not require longsome digestion or microscopy by trained personnel.
RESUMO
The parasitic roundworm Trichinella spiralis is most commonly transmitted to humans through consumption of raw or undercooked meat of infected pigs or game. To prevent human infection, slaughterhouses perform meat safety surveillance using the gold standard "Magnetic Stirrer Method". We introduce a fast and objective method using automated detection of specific Trichinella spiralis antigens by a newly developed immunoassay based on chemiluminescence (ChLIA). Panel A comprised muscle tissue samples from non-infected pigs (n=37). Panel B comprised muscle tissue samples from non-infected pigs spiked with different amounts of Trichinella larvae without collagen capsules (n=56). Panel C contained muscle tissue samples from experimentally infected pigs including Trichinella larvae encapsulated in collagen (n=32). Each sample was shredded with PBS buffer in a knife mill, destroying Trichinella larvae. Following centrifugation, the supernatant (muscle tissue extract containing released excretory and secretory Trichinella spiralis antigens) was used for Trichinella-specific antigen detection by the new Trichinella ChLIA. The overall accuracy of the Trichinella ChLIA was 97.6 %. The specificity of the Trichinella ChLIA was 100 % (panel A). The sensitivity in samples from experimentally infected pigs was 100 % representing a detection limit of 0.01 larvae per gram. Cross-reactivity with parasites other than Trichinella spp. was not observed. This new meat inspection method for the detection of Trichinella spiralis antigens presents high specificity and high sensitivity, especially in truly infected samples. In contrast to the gold standard, this new approach to meat safety surveillance does not require longsome digestion or microscopy by trained personnel.
RESUMO
Myopathies are a rare type of acquired, chronic autoimmune diseases of the skeletal muscles and affect both children and adults. The hallmark symptoms of idiopathic inflammatory myopathies (IIM) are muscle inflammation, proximal muscle weakness and disability, arthritis, cutaneous rashes, calcinosis, ulceration, malignancy and interstitial lung disease (ILD). Subforms of IIM include polymyositis, dermatomyositis, cancer-related myositis and sporadic inclusion body myositis. Autoantibodies function as biomarkers for diagnosis of IIM and can be used to delimit clinically distinguishable IIM subforms. To maximise the diagnostic information it is essential to perform comprehensive multiparametric serological testing including both screening and confirmation tests.