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1.
Methods Mol Biol ; 2578: 199-208, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36152289

RESUMEN

Flavivirus are the most alarming prevalent viruses worldwide due to its vast impact on public health. Most early symptoms of diseases caused by Flavivirus are similar among each other and to other febrile illnesses making the clinical differential diagnosis challenging. In addition, due to cross-reactivity and a relatively limited persistence of viral RNA in infected individuals, the current available diagnosis strategies fail to efficiently provide a differential viral identification. In this context, virus-specific tests are essential to improve patient care, as well as to facilitate disease surveillance and the effective control of transmission. Here, we describe the use of protein microarrays as an effective tool for screening peptides differentially recognized by anti-Yellow Fever virus antibodies induced by vaccination or by natural viral infection.


Asunto(s)
Flavivirus , Anticuerpos Antivirales , Reacciones Cruzadas , Flavivirus/genética , Humanos , Péptidos , ARN Viral/genética
2.
Methods Mol Biol ; 2578: 209-217, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36152290

RESUMEN

In SARS-CoV-2 pandemic scenario, the identification of rapid methods to detect antibodies against coronavirus has been a wide and urgent issue. Epitope mapping on peptide microarrays is a rapid way to identify sequences with a high immunoreactivity. The process begins with a proteome-wide screening, based on immune affinity; the use of a high-density microarray is followed by a validation phase, where a restricted panel of probes is tested using peptide microarrays; peptide sequences are immobilized through a click-based strategy.COVID-19-positive sera are tested and immuno-domains regions are identified on SARS-CoV-2 spike (S), nucleocapsid (N) protein, and Orf1ab polyprotein. An epitope on N protein (region 155-171) provided good diagnostic performance in discriminating COVID-19-positive vs. healthy individuals. Using this sequence, 92% sensitivity and 100% specificity are reached for IgG detection in COVID-19 samples, and no cross-reactivity with common cold coronaviruses is detected. Overall, epitope 155-171 from N protein represents a promising candidate for further development and rapid implementation in serological tests.


Asunto(s)
COVID-19 , SARS-CoV-2 , Anticuerpos Antivirales , COVID-19/diagnóstico , Mapeo Epitopo , Epítopos , Humanos , Inmunidad , Inmunoglobulina G , Poliproteínas , Proteoma , SARS-CoV-2/genética , Glicoproteína de la Espiga del Coronavirus
3.
Arch Argent Pediatr ; 121(1): e202202595, 2023 02 01.
Artículo en Inglés, Español | MEDLINE | ID: mdl-35984671

RESUMEN

Introduction. In Argentina, health care workers have been the first ones to receive the COVID-19 vaccine, but there are still few data on the production of anti-S IgG antibodies. Objectives. To assess specific IgG against the SARS-CoV-2 spike protein (anti-S IgG) after the vaccination of health care workers from a children's hospital. To explore the association between the presence of these antibodies, age, and history of prior infection. Population and methods. Cross-sectional study in 193 workers who received both doses of the two- component Sputnik V vaccine. The anti-S IgG antibody titer was measured and age, history of prior SARS-CoV-2 infection, and date of vaccination were recorded. Results. Anti-S IgG antibodies were produced in 98.6% of the subjects. The titer was higher in those with prior infection (p < 0.001), but no relationship was established with subjects' age. Conclusion. We provide data on post-vaccination production of IgG anti-S antibodies among health care workers from a children's hospital and explore some predictors.


Introducción. En Argentina, el personal de salud ha sido el primero en vacunarse contra COVID-19, pero todavía existen pocos datos sobre la producción de anticuerpos IgG anti-S. Objetivos. Evaluar IgG específica contra glicoproteína spike del SARS-CoV-2 (IgG anti-S) posvacunación en personal de un hospital pediátrico. Explorar la asociación entre presencia de dichos anticuerpos, edad y antecedente de infección previa. Población y métodos. Estudio transversal que incluyó 193 trabajadores vacunados con los dos componentes de la vacuna Sputnik V. Se pesquisó el título de IgG anti-S y se registraron edad, antecedente de infección previa por SARS-CoV-2 y fecha de la vacunación. Resultados. El 98,6 % de los sujetos generó IgG anti-S. El título fue mayor en quienes habían cursado infección previamente (p <0,001), pero no hubo relación con la edad de los sujetos. Conclusión. Aportamos datos de generación de anticuerpos IgG anti-S posvacunación en personal de salud de un hospital pediátrico y exploramos algunos predictores.


Asunto(s)
COVID-19 , Personal de Salud , SARS-CoV-2 , Anticuerpos Antivirales , COVID-19/inmunología , Vacunas contra la COVID-19 , Estudios Transversales , Hospitales Pediátricos , Humanos , Inmunoglobulina G , SARS-CoV-2/inmunología , Glicoproteína de la Espiga del Coronavirus
5.
Talanta ; 251: 123783, 2023 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-35977451

RESUMEN

The current approaches of diagnostic platforms for detecting SARS-CoV-2 infections mostly relied on adapting the existing technology. In this work, a simple and low-cost electrochemical sensing platform for detecting SAR-CoV-2 antigen was established. The proposed sensor combined the innovative disposable paper-based immunosensor and cost-effective plant-based anti-SARS-CoV-2 monoclonal antibody CR3022, expressed in Nicotiana benthamiana. The cellulose nanocrystal was modified on screen-printed graphene electrode to provide the abundant COOH functional groups on electrode surface, leading to the high ability for antibody immobilization. The quantification of the presence receptor binding domain (RBD) spike protein of SARS-CoV-2 was performed using differential pulse voltammetry by monitoring the changing current of [Fe(CN)6]3-/4- redox solution. The current change of [Fe(CN)6]3-/4- before and after the presence of target RBD could be clearly distinguished, providing a linear relationship with RBD concentration in the range from 0.1 pg/mL to 500 ng/mL with the minimum limit of detection of 2.0 fg/mL. The proposed platform was successfully applied to detect RBD in nasopharyngeal swab samples with satisfactory results. Furthermore, the paper-based immunosensor was extended to quantify the RBD level in spiked saliva samples, demonstrating the broadly applicability of this system. This electrochemical paper-based immunosensor has the potential to be employed as a point-of-care testing for COVID-19 diagnosis.


Asunto(s)
Técnicas Biosensibles , COVID-19 , Grafito , Anticuerpos Monoclonales/química , Anticuerpos Neutralizantes , Anticuerpos Antivirales , Técnicas Biosensibles/métodos , COVID-19/diagnóstico , Prueba de COVID-19 , Celulosa , Técnicas Electroquímicas/métodos , Grafito/química , Humanos , Inmunoensayo/métodos , SARS-CoV-2 , Glicoproteína de la Espiga del Coronavirus
6.
Methods Mol Biol ; 2585: 15-21, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36331761

RESUMEN

Immunostained plaque assay based on the specific antibody binding to viral antigen enables the detection and titration of virus infectivity, especially for viruses that could not form plaques using the classical crystal violet or neutral red staining methods. Here we describe the application of this method to quantify viral titers of wild-type West Nile virus (WNV-WT) and replication-defective WNV-ΔNS1 virus.


Asunto(s)
Fiebre del Nilo Occidental , Virus del Nilo Occidental , Humanos , Carga Viral , Replicación Viral , Pruebas Serológicas , Anticuerpos Antivirales , Ensayo de Placa Viral
7.
Methods Mol Biol ; 2585: 119-125, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36331770

RESUMEN

West Nile virus (WNV) is one of the leading causes of arboviral encephalitis in the United States but is often underdiagnosed. Despite the wide breadth of WNV-induced clinical disease syndromes, many of the symptoms associated with WNV are nonspecific at the time of presentation; thus, choosing the right diagnostic tool is essential to not only understand the true burden of disease but also provide pathogen-directed interventions for WNV-infected patients. In this chapter, we briefly discuss the three most common types of diagnostic methods for WNV in human clinical samples: nucleic acid detection, enzyme-linked immunoassay (ELISA), and plaque reduction neutralization test (PRNT) and present the method for PRNT.


Asunto(s)
Fiebre del Nilo Occidental , Virus del Nilo Occidental , Humanos , Anticuerpos Antivirales , Ensayo de Inmunoadsorción Enzimática/métodos , Fiebre del Nilo Occidental/diagnóstico
8.
J Theor Biol ; 556: 111280, 2023 Jan 07.
Artículo en Inglés | MEDLINE | ID: mdl-36202234

RESUMEN

Compelling evidence continues to build to support the idea that SARS-CoV-2 Neutralizing Antibody (NAb) levels in an individual can serve as an important indicator of the strength of protective immunity against infection. It is not well understood why NAb levels in some individuals remain high over time, while in others levels decline rapidly. In this work, we present a two-state mathematical model of within-host NAb dynamics in response to vaccination. By fitting only four host-specific parameters, the model is able to capture individual-specific NAb levels over time as measured by the AditxtScore™ for NAbs. The model can serve as a foundation for predicting NAb levels in the long-term, understanding connections between NAb levels, protective immunity, and breakthrough infections, and potentially guiding decisions about whether and when a booster vaccination may be warranted.


Asunto(s)
COVID-19 , SARS-CoV-2 , Humanos , COVID-19/prevención & control , Vacunas contra la COVID-19 , Anticuerpos Antivirales , Vacunación , Anticuerpos Neutralizantes , Modelos Teóricos
9.
Acta Trop ; 237: 106736, 2023 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-36374844

RESUMEN

Genetic and phylogenetic studies indicated that Zika virus (ZIKV) has evolved into 2 major lineages, the African and Asian. However, ZIKV has been described as a single serotype. This study aimed at assessing the cross-neutralization between ZIKV African and Asian lineages strains. Sixthy-five samples collected in 2007 and 30 samples collected from the same subjects in 2011/2012 in West Africa and positive to neutralizing antibody against ZIKV MR-766 strain (African lineage) were tested against ZIKV H/PF/2013 strain (Asian lineage) by microneutralization assay. All samples showing neutralizing antibodies against MR-766 strain showed also neutralizing activity against H/PF/2013 strain, although with lower titers. This is consistent with about 120 amino acid differences between the two strains. Despite differences in the magnitude of neutralizing activity against different ZIKV strains, all samples showed neutralizing antibody titers considered to be protective.


Asunto(s)
Infección por el Virus Zika , Virus Zika , Humanos , Virus Zika/genética , Sueros Inmunes , Filogenia , Anticuerpos Antivirales , Anticuerpos Neutralizantes
10.
J Theor Biol ; 557: 111334, 2023 Jan 21.
Artículo en Inglés | MEDLINE | ID: mdl-36306828

RESUMEN

The COVID-19 pandemic has underscored the need to understand the dynamics of SARS-CoV-2 respiratory infection and protection provided by the immune response. SARS-CoV-2 infections are characterized by a particularly high viral load, and further by the small number of inhaled virions sufficient to generate a high viral titer in the nasal passage a few days after exposure. SARS-CoV-2 specific antibodies (Ab), induced from vaccines, previous infection, or inhaled monoclonal Ab, have proven effective against SARS-CoV-2 infection. Our goal in this work is to model the protective mechanisms that Ab can provide and to assess the degree of protection from individual and combined mechanisms at different locations in the respiratory tract. Neutralization, in which Ab bind to virion spikes and inhibit them from binding to and infecting target cells, is one widely reported protective mechanism. A second mechanism of Ab protection is muco-trapping, in which Ab crosslink virions to domains on mucin polymers, effectively immobilizing them in the mucus layer. When muco-trapped, the continuous clearance of the mucus barrier by coordinated ciliary propulsion entrains the trapped viral load toward the esophagus to be swallowed. We model and simulate the protection provided by either and both mechanisms at different locations in the respiratory tract, parametrized by the Ab titer and binding-unbinding rates of Ab to viral spikes and mucin domains. Our results illustrate limits in the degree of protection by neutralizing Ab alone, the powerful protection afforded by muco-trapping Ab, and the potential for dual protection by muco-trapping and neutralizing Ab to arrest a SARS-CoV-2 infection. This manuscript was submitted as part of a theme issue on "Modelling COVID-19 and Preparedness for Future Pandemics".


Asunto(s)
COVID-19 , SARS-CoV-2 , Humanos , Pandemias , Anticuerpos Antivirales , Sistema Respiratorio , Mucinas
11.
Rev Med Inst Mex Seguro Soc ; 61(Suppl 1): 28-32, 2023 01 01.
Artículo en Español | MEDLINE | ID: mdl-36378067

RESUMEN

Background: COVID-19 pandemic spread around the world swiftly; there are several diagnostic strategies available. Health workers, especially medical residents (MR), are a high-risk population for acquiring this infection. Objective: To estimate the seroprevalence of antibodies against SARS-CoV-2 and the associated factors in MR of a third level hospital. Material and methods: 330 MR from different specialties were evaluated with a questionnaire and collection of blood samples for analysis by microparticle chemiluminescent immunoassay. The prevalence of previous infection was defined by seropositivity of these antibodies. Descriptive statistics and concordance between the RT-PCR tests and the presence of anti-SARS-CoV-2 IgG were used. Results: Of 330 MR, 84.5% actively participated in COVID patient care. One out of 3 reported symptoms of COVID-19; in 67.6% the possible site of infection was a hospital setting not associated with the COVID area. Out of 71 symptomatic subjects, 61.9% underwent RT-PCR against SARS-CoV-2; 20 were positive. In 15.8% of the total, the presence of anti-SARS-CoV-2 IgG antibodies was determined. Only 1 out of 3 subjects with a positive PCR had antibodies, and 11.3% of the cases, even with a positive RT-PCR test, did not develop humoral immunity. Conclusions: The seroprevalence was lower than that reported at the national level, potentially due to protection measures. The main risk factor was contact with the virus in areas of the hospital not related to COVID, making it imperative to reinforce security protocols in those spaces.


Introducción: la pandemia por COVID-19 se extendió rápidamente a nivel mundial; hay disponibles varias estrategias de diagnóstico. Los trabajadores de la salud, en especial los médicos residentes (MR), son una población de alto riesgo para adquirir dicha infección. Objetivo: estimar la seroprevalencia de anticuerpos contra el SARS-CoV-2 y los factores asociados en los MR de un hospital de tercer nivel. Material y métodos: se evaluaron 330 MR de diferentes especialidades con un cuestionario y recolección de muestras de sangre para análisis mediante un inmunoensayo quimioluminiscente de micropartículas. La prevalencia de infección previa se definió por seropositividad de estos anticuerpos. Se utilizó estadística descriptiva y concordancia entre las pruebas RT-PCR y presencia de IgG anti-SARS-CoV-2. Resultados: de los 330 MR, 84.5% participó activamente en atención de pacientes COVID. Uno de cada tres refirió síntomas de COVID-19; 67.6% tuvo posible sitio de contagio en ámbito hospitalario no asociado a Área COVID. De los 71 sujetos sintomáticos, 61.9% se realizaron RT-PCR; 20 fueron positivas. En 15.8% del total se determinó la presencia de anticuerpos IgG anti-SARS-CoV-2. Solo uno de cada tres sujetos con PCR positiva presentó anticuerpos y 11.3% de los casos, aun con RT-PCR positiva, no desarrolló inmunidad humoral. Conclusiones: la seroprevalencia fue menor que la reportada a nivel nacional, potencialmente por medidas de protección. El principal factor de riesgo fue el contacto con el virus en áreas del hospital no relacionadas a COVID, por lo que es imperativo reforzar los protocolos de seguridad en esos espacios.


Asunto(s)
COVID-19 , Médicos , Humanos , Estudios Seroepidemiológicos , SARS-CoV-2 , Pandemias , COVID-19/diagnóstico , COVID-19/epidemiología , Anticuerpos Antivirales , Inmunoglobulina G , Personal de Salud
12.
J Infect Chemother ; 29(1): 61-66, 2023 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-36152928

RESUMEN

BACKGROUND: Data are limited regarding the safety of and antibody response to the BNT162b2 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) messenger ribonucleic acid vaccine in adolescents and young adults with underlying disease. METHODS: This prospective observational study enrolled patients age 12-25 years with chronic underlying disease who received 2 doses of BNT162b2. A 18-item questionnaire was used to assess adverse events within 7 days post-vaccination, and data regarding severe adverse events were collected from electronic medical records. An antibody titer for the receptor-binding domain of the spike protein in SARS-CoV-2 was used to assess antibody response after the second vaccine dose. RESULTS: Study participants were 429 patients (241 [56.2%] age 12-15 years; 188 [43.8%] age 16-25 years). The most common underlying diseases were genetic or chromosomal abnormalities and/or congenital anomalies, followed by endocrine or metabolic diseases; 32% of participants were immunocompromised. Severe adverse events were observed after the second dose in 1 (0.4%) patient age 12-15 years and in 2 (1.1%) patients age 16-25 years; all patients recovered. Seropositivity after the second vaccine dose was 99.0%. The geometric mean antibody titer was higher in patients age 12-15 years versus 16-25 years (1603.3 [1321.8-1944.7] U/mL vs. 949.4 [744.2-1211.0] U/mL). Compared with immunocompetent patients, immunocompromised patients had a lower antibody titer (2106.8 [1917.5-2314.7] U/mL vs. 467.9 [324.4-674.8] U/mL). CONCLUSIONS: Vaccination with BNT162b2 was acceptably safe and immunogenic for adolescents and young adults with underlying disease.


Asunto(s)
Vacunas contra la COVID-19 , COVID-19 , SARS-CoV-2 , Adolescente , Adulto , Niño , Humanos , Adulto Joven , Anticuerpos Antivirales , Formación de Anticuerpos , Vacuna BNT162 , COVID-19/prevención & control , Vacunas contra la COVID-19/efectos adversos
13.
J Exp Med ; 220(1)2023 Jan 02.
Artículo en Inglés | MEDLINE | ID: mdl-36342455

RESUMEN

Inborn and acquired deficits of type I interferon (IFN) immunity predispose to life-threatening COVID-19 pneumonia. We longitudinally profiled the B cell response to mRNA vaccination in SARS-CoV-2 naive patients with inherited TLR7, IRF7, or IFNAR1 deficiency, as well as young patients with autoantibodies neutralizing type I IFNs due to autoimmune polyendocrine syndrome type-1 (APS-1) and older individuals with age-associated autoantibodies to type I IFNs. The receptor-binding domain spike protein (RBD)-specific memory B cell response in all patients was quantitatively and qualitatively similar to healthy donors. Sustained germinal center responses led to accumulation of somatic hypermutations in immunoglobulin heavy chain genes. The amplitude and duration of, and viral neutralization by, RBD-specific IgG serological response were also largely unaffected by TLR7, IRF7, or IFNAR1 deficiencies up to 7 mo after vaccination in all patients. These results suggest that induction of type I IFN is not required for efficient generation of a humoral response against SARS-CoV-2 by mRNA vaccines.


Asunto(s)
Linfocitos B , Vacunas contra la COVID-19 , COVID-19 , Interferón Tipo I , Humanos , Anticuerpos Neutralizantes , Anticuerpos Antivirales , Autoanticuerpos , COVID-19/inmunología , COVID-19/prevención & control , SARS-CoV-2 , Glicoproteína de la Espiga del Coronavirus/genética , Receptor Toll-Like 7/genética , Vacunación , Vacunas de ARNm , Vacunas contra la COVID-19/inmunología , Linfocitos B/inmunología , Interferón Tipo I/deficiencia
14.
Life Sci Alliance ; 6(1)2023 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-36344085

RESUMEN

The efficacy of the current influenza vaccines is frequently reduced because of antigenic drift, a trade-off of developing improved vaccines with broad cross-protective activity against influenza A viruses. In this study, we have successfully constructed a chimeric cytokine (CC) comprising the M2 protein, influenza A neuraminidase stalk, and interleukin-12. We produced virus-like particles (VLPs) containing CC and influenza hemagglutinin (HA) proteins using a baculovirus system in Eri silkworm pupae. The protective efficacy of the CCHA-VLP vaccine was evaluated in mice. The CCFkH5HA-VLP vaccine increased the survival rates of BALB/c mice, infected with a lethal dose of PRH1 and HKH5 viruses, to 80% and 100%, respectively. The results suggested that CCHA-VLP successfully induced potent cross-reactive protective immunity against infection with homologous and heterologous subtypes of the influenza A virus. This is the first study to design a CC-containing HA-VLP vaccine and validate its protective efficacy.


Asunto(s)
Vacunas contra la Influenza , Gripe Humana , Infecciones por Orthomyxoviridae , Vacunas de Partículas Similares a Virus , Ratones , Animales , Humanos , Gripe Humana/prevención & control , Vacunas de Partículas Similares a Virus/genética , Citocinas , Infecciones por Orthomyxoviridae/prevención & control , Anticuerpos Antivirales , Hemaglutininas , Ratones Endogámicos BALB C
15.
J Infect Chemother ; 29(1): 112-114, 2023 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-36167304

RESUMEN

Vaccines having aided in escaping the majority of the population from immunological naïvety, our strategies are now shifting towards an increased focus on identifying and protecting the extremely vulnerable. We here describe the results of testing 12 patients, those with lymphoid malignancies having been targeted their B-cells for therapy with rituximab-containing regimens or a Bruton tyrosine kinase inhibitor, for anti-SARS-CoV-2 spike antibodies after receiving the BNT162b2 mRNA vaccine doses. The interval from last dosing of B-cell depletion therapy to SARS-CoV-2 vaccination was at median 5.3 (range 3.1-6.6) months. Using the 'seroprotection' threshold of 775 [BAU/mL] for the anti-spike antibody titer, our finding points out the crucial unresponsiveness of the targeted population with 0/12 (0%) achieving 'seroprotection'. Although IgG seroconversion was observed in 4/12 (33%), supporting the overall benefit of vaccination, the figures still point out a potential need for optimization of practice. IgA was further less responsive (unsuccessful 'seroconversion' in 11/12 (92%)), implicating an underlying class switch defect. Those with depletion on B-cells are caught at a dilemma between, being too early and too late on receiving SARS-CoV-2 vaccines. They wish to get over their immunological naïvety at the earliest, while, in order to assure quality immune memory, are also required to hold the patience for their B-cells to repopulate. Although it remains an issue whether intensified vaccine schedules and/or regimens will lead to stronger immunogenicity or more effective boosters for non-responders, we shall take advantage of every increasing evidence in order to optimize current options.


Asunto(s)
COVID-19 , Neoplasias , Vacunas Virales , Humanos , Formación de Anticuerpos , Vacunas contra la COVID-19 , Vacuna BNT162 , Cambio de Clase de Inmunoglobulina , Vacunas Virales/farmacología , COVID-19/prevención & control , SARS-CoV-2 , Vacunación , Anticuerpos Antivirales
16.
J Infect Chemother ; 29(1): 39-42, 2023 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-36168999

RESUMEN

BACKGROUND: To mitigate the COVID-19 pandemic, many countries have recommended the use of booster vaccinations. The relationship between the degree of adverse vaccine reactions and elevated antibody titers is of interest; however, no studies have investigated the temporal changes in antibody titers based on repeated measurements after a third dose of the BNT162b2 vaccine. METHODS: This prospective longitudinal cohort study was conducted with 62 healthcare workers who received a third dose of the BNT162b2 at Okayama University Hospital, Japan. Venous blood draw and fingertip whole blood test sample collection were conducted at the early (3-13 days) and 1-month time points; only FWT sample collection was conducted at the 2-month time point. Information on adverse reactions within 1 week after vaccination was also obtained. The association between fever of 37.5 °C or higher and antibody titers after the third dose of BNT162b2 was examined using a mixed-effects model and Poisson regression with robust variance. RESULTS: A trend toward higher antibody titers in the early period after vaccination was observed in the febrile individuals, but the differences were not significant at 1 and 2 months post-vaccination (the partial regression coefficient for fever was 8094.3 [-1910.2, 18,098.8] at 1 month after vaccination, and 1764.1 [-4133.9, 7662.1] at 2 months after vaccination in the adjusted models). CONCLUSION: The findings suggest that the presence of fever after the third vaccine does not predict a sustained elevation in serum antibody titers.


Asunto(s)
COVID-19 , Vacunas , Humanos , Vacuna BNT162 , Estudios Prospectivos , Estudios Longitudinales , Pandemias , COVID-19/prevención & control , Anticuerpos Antivirales
17.
J Virol Methods ; 311: 114643, 2023 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-36332715

RESUMEN

Goose astrovirus (GAstV) characterized by articular and visceral gout, is an emerging pathogen with a wide distribution on mainland China, leading to serious economic losses in the goose-raising industry. Because vaccines to prevent GAstV infections are not available currently, early diagnosis is critical when treating symptomatic geese and in preventing GAstV transmission. In this context, a highly sensitive indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) based on a monoclonal antibody (mAb) to detect GAstV-specific antibodies from geese was developed, and the detections were optimized. A series of experiments proved that the ic-ELISA shows excellent diagnostic performance and discriminatory power with high sensitivity and specificity. The ic-ELISA for GAstV detection was applied on 67 field serum samples, and comparing the detection results with the virus neutralization test verified the accuracy of the ic-ELISA. The correlation coefficient between the ic-ELISA and the virus neutralization test was 80%, demonstrating the proposed ic-ELISA method could be a useful and effective tool for the diagnostic, serological epidemiological investigation and immune monitoring of the GAstV in goose-producing regions.


Asunto(s)
Infecciones por Astroviridae , Avastrovirus , Animales , Gansos , Anticuerpos Antivirales , Ensayo de Inmunoadsorción Enzimática/métodos , Infecciones por Astroviridae/diagnóstico , Infecciones por Astroviridae/veterinaria
18.
J Virol Methods ; 311: 114644, 2023 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-36332713

RESUMEN

Bovine leukemia virus (BLV) is an enveloped virus, found worldwide that can infect cattle and induce many subclinical symptoms and malignant tumors. BLV infection causes severe economic losses in the cattle industry. The identification of BLV-infected cattle for segregation or elimination would be the most effective way to halt the spread of BLV infection on farms, owing to the lack of effective treatments and vaccines. Therefore, antibody detection against the viral glycoprotein gp51 is an effective method for diagnosing BLV-infected animals. In this study, ten different subregions of gp51 containing a common B cell epitope are vital for developing antigens as epitope-driven vaccine design and immunological assays. Such antigens were produced in Escherichia coli expression system to react with antibodies in the serum from BLV-infected cattle and compete for antigenicity. Recombinant His-gp5156-110 and gp5133-301(full) had the same sensitivity in BLV-positive sera, indicating that antibodies responded to the limited subregion of viral gp51, a common B cell epitope. This finding provides significant information for antigen selection in BLV to use in antibody detection assays. Further studies are needed to evaluate the antigenicity of His-gp5156-110 and gp5133-301(full) as antigens for antibody detection assays using a larger number of bovine serum samples.


Asunto(s)
Infecciones por Deltaretrovirus , Leucosis Bovina Enzoótica , Virus de la Leucemia Bovina , Animales , Bovinos , Virus de la Leucemia Bovina/genética , Proteínas del Envoltorio Viral , Epítopos de Linfocito B/metabolismo , Anticuerpos Antivirales , Leucosis Bovina Enzoótica/diagnóstico
19.
Anal Biochem ; 660: 114929, 2023 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-36270332

RESUMEN

Detection and quantification of antibodies, especially immunoglobulin G (IgG), is a cornerstone of ELISAs, many diagnostics, and the development of antibody-based drugs. Current state-of-the-art immunoassay techniques for antibody detection require species-specific secondary antibodies and carefully-controlled bioconjugations. Poor conjugation efficiency degrades assay performance and increases the risk of clinical false positives due to non-specific binding. We developed a generic, highly-sensitive platform for IgG quantification by fusing the IgG-Fc binding Z domain of Staphylococcal Protein A with the ultrabright bioluminescence reporter Nanoluc-luciferase (Nluc). We demonstrated the application of this fusion protein in a sandwich IgG detection immunoassay using surface-bound antigens to capture target IgG and protein A-Nanoluc fusion as the detector. We optimized the platform's sensitivity by incorporating multiple repeats of the Z domain into the fusion protein constructs. Using rabbit and mouse anti-SARS-CoV-2 Nucleoprotein IgGs as model analytes, we performed ELISAs in two different formats, either with SARS-CoV-2 Nucleoprotein as the capture antigen or with polyclonal chicken IgY as the capture antibody. Using standard laboratory equipment, the platform enabled the quantitation of antibody analytes at concentrations as low as 10 pg/mL (67 fM).


Asunto(s)
COVID-19 , Inmunoglobulina G , Ratones , Conejos , Animales , Proteína Estafilocócica A , SARS-CoV-2 , Anticuerpos Antivirales , Inmunoensayo/métodos , Nucleoproteínas , Sensibilidad y Especificidad
20.
J Virol Methods ; 311: 114641, 2023 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-36328082

RESUMEN

Antibody-dependent enhancement (ADE) is one of the pathogenic mechanisms related to disease severity in dengue virus infection. Conventional assays for detecting ADE activity usually require several days. In this study, we established a rapid assay system to evaluate ADE activity in dengue-seropositive samples using single round infectious particles (SRIPs). Human Fc-gamma receptor-bearing cells (K562 and Mylc cells) were infected with SRIP antigen in the presence of human serum samples to measure ADE activity. Two assay protocols were introduced: (i) rapid assay with 5 h of incubation, and (ii) semi-rapid assay with 24 h of incubation. The rapid assay requires a large quantity of SRIP antigen and gives results in half a day. Although the semi-rapid assay requires slightly more than a day, it can be performed using only a small amount of SRIP. Interestingly, the range of the number of Mylc cells required for the semi-rapid assay was wider than that of K562 cells. Significant correlations were observed between the rapid and semi-rapid assays for both cell types. Although it is difficult to judge which protocol best reflects the current immune status in vivo, both assays could rapidly provide valuable information regarding the risk assessment for severe diseases.


Asunto(s)
Virus del Dengue , Dengue , Virosis , Humanos , Acrecentamiento Dependiente de Anticuerpo , Anticuerpos Antivirales
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