RESUMO
In 2022 the World Health Organization declared a Public Health Emergency for an outbreak of mpox, the zoonotic Orthopoxvirus (OPV) affecting at least 104 nonendemic locations worldwide. Serologic detection of mpox infection is problematic, however, due to considerable antigenic and serologic cross-reactivity among OPVs and smallpox-vaccinated individuals. In this report, we developed a high-throughput multiplex microsphere immunoassay using a combination of mpox-specific peptides and cross-reactive OPV proteins that results in the specific serologic detection of mpox infection with 93% sensitivity and 98% specificity. The New York State Non-Vaccinia Orthopoxvirus Microsphere Immunoassay is an important tool to detect subclinical mpox infection and understand the extent of mpox spread in the community through retrospective analysis.
Assuntos
Mpox , Orthopoxvirus , Humanos , Estudos Retrospectivos , Infecções Assintomáticas , Bioensaio , Reações CruzadasRESUMO
Passive transfer of antibodies from COVID-19 convalescent patients is being used as an experimental treatment for eligible patients with SARS-CoV-2 infections. The United States Food and Drug Administration's (FDA) guidelines for convalescent plasma initially recommended target antibody titers of 160. We evaluated SARS-CoV-2 neutralizing antibodies in sera from recovered COVID-19 patients using plaque reduction neutralization tests (PRNT) at moderate (PRNT50) and high (PRNT90) stringency thresholds. We found that neutralizing activity significantly increased with time post symptom onset (PSO), reaching a peak at 31-35 days PSO. At this point, the number of sera having neutralizing titers of at least 160 was approximately 93% (PRNT50) and approximately 54% (PRNT90). Sera with high SARS-CoV-2 antibody levels (>960 enzyme-linked immunosorbent assay titers) showed maximal activity, but not all high-titer sera contained neutralizing antibody at FDA recommended levels, particularly at high stringency. These results underscore the value of serum characterization for neutralization activity.
Assuntos
Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , COVID-19/terapia , Testes de Neutralização , Ensaio de Imunoadsorção Enzimática , Humanos , Imunização Passiva , Soroterapia para COVID-19RESUMO
The recent outbreak of Zika virus (ZIKV) in the Americas has challenged diagnostic laboratory testing strategies. At the Wadsworth Center, ZIKV serological testing was performed for over 10,000 specimens, using a combination of an enzyme-linked immunosorbent assay (ELISA) for IgM antibodies (Abs) to ZIKV, a polyvalent microsphere immunoassay (MIA) to detect Abs broadly reactive with flaviviruses, and a plaque reduction neutralization test (PRNT) for further testing. Overall, 42% of patients showed serological evidence of flavivirus infection (primarily past dengue virus [DENV] infection), while 7% possessed IgM Abs to ZIKV and/or DENV. ZIKV IgM Abs typically arose within 3 to 4 days, with only one instance of duration beyond 100 days after reported symptoms. PRNT analysis of 826 IgM-positive specimens showed 7% positive neutralization to ZIKV alone, 9% to DENV alone, and 85% to both ZIKV and DENV. Thus, the extensive Ab cross-reactivity among flaviviruses significantly reduced the value of performing PRNT analysis, especially when a traditional paired serum algorithm with viral neutralization titering was used. Nevertheless, the finding of a negative ZIKV result by PRNT was invaluable for reassuring both physicians and patients. The MIA detected both IgM and IgG, which enabled us to identify patients who presented without IgM anti-ZIKV Abs but still had ZIKV-specific neutralizing Abs. On the basis of these results, a new algorithm, which included an IgM Ab capture (MAC)-ELISA to detect recent infection, a flavivirus MIA to identify patients no longer producing IgM, and a single-dilution PRNT for ZIKV exclusion and occasional discrimination of ZIKV and DENV, was implemented.
Assuntos
Testes Sorológicos/métodos , Infecção por Zika virus/diagnóstico , Zika virus/imunologia , Algoritmos , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Reações Cruzadas , Vírus da Dengue/imunologia , Humanos , Imunoensaio , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Testes de Neutralização , New York , Guias de Prática Clínica como Assunto , Testes Sorológicos/tendências , Zika virus/isolamento & purificaçãoRESUMO
West Nile virus (WNV) neuroinvasive disease (WNND) occurs in approximately 1 percent of WNV-infected patients and typically presents as encephalitis, meningitis, or acute flaccid paralysis (AFP). WNND remains a difficult inpatient diagnosis, creating significant challenges for prognostication and therapy selection. We characterized the clinical and diagnostic features of WNND cases at two major academic medical centers in New York City in routine clinical practice. We retrospectively reviewed the charts of thirty-six patients with WNND, including twenty-six encephalitis, four meningitis, and six AFP cases. The most common presenting symptoms were fever (86.1%) and gastrointestinal symptoms (38.9%) in addition to altered mental status (72.2%), lethargy (63.9%), gait disturbances (46.2%), and headache (44.4%). Fourteen (48.3%) patients displayed acute magnetic resonance imaging (MRI) findings, particularly T2 hyperintensities in the bilateral thalami, brainstem, and deep white matter. New York State Department of Health WNV CSF IgM testing was utilized for diagnosis in 58.3% of patients; however, just 38.1% had the result by discharge, compared to 85.6% of those who underwent serum IgM testing. The median length of stay was 13.5 days, 38.9% were intubated, and three patients (8.9%) died during acute hospitalization. Our findings underscore the morbidity, mortality, and diagnostic challenges of WNND, suggesting the potential utility of serum IgM testing in combination with confirmatory CSF testing to expedite diagnosis in the acute setting.
RESUMO
In 2022 the World Health Organization declared a Public Health Emergency for an outbreak of mpox, the zoonotic Orthopoxvirus (OPV) affecting at least 103 non-endemic locations world-wide. Serologic detection of mpox infection is problematic, however, due to considerable antigenic and serologic cross-reactivity among OPVs and smallpox-vaccinated individuals. In this report, we developed a high-throughput multiplex microsphere immunoassay (MIA) using a combination of mpox-specific peptides and cross-reactive OPV proteins that results in the specific serologic detection of mpox infection with 93% sensitivity and 98% specificity. The New York State Non-Vaccinia Orthopoxvirus Microsphere Immunoassay is an important diagnostic tool to detect subclinical mpox infection and understand the extent of mpox spread in the community through retrospective analysis.
RESUMO
Although measles was eliminated in the United States in 2000, a severe outbreak occurred between October 2018 and September 2019. New York was especially hard hit. Serology played an integral role in determining immune status (IgG) and identifying, along with molecular analyses, acute measles infections (IgM). Although an indirect immunofluorescence assay (IFA) was historically used by the New York State Department of Health for measles IgM detection, a higher throughput assay was needed to address the increased specimen numbers. Four commercial enzyme-linked immunosorbent assays (ELISAs) were evaluated for sensitivity and specificity in detecting measles IgM. Two ELISA formats were compared, indirect ELISA and IgM antibody capture. Both formats had comparable specificity as determined by cross-reactivity to non-measles specimens. Overall, the sensitivity of the capture ELISAs was greater than the indirect ELISAs and comparable to the indirect immunofluorescence assay with benefits regarding capacity, cost, and turnaround time.
Assuntos
Anticorpos Antivirais , Sarampo , Surtos de Doenças , Ensaio de Imunoadsorção Enzimática , Humanos , Imunoglobulina M , Sarampo/diagnóstico , Sarampo/epidemiologia , New York/epidemiologia , Sensibilidade e Especificidade , Testes SorológicosRESUMO
Coronavirus disease 2019 (COVID-19) is associated with a wide spectrum of disease presentation, ranging from asymptomatic infection to acute respiratory distress syndrome (ARDS). Paradoxically, a direct relationship has been suggested between COVID-19 disease severity and the levels of circulating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific antibodies, including virus-neutralizing titers. A serological analysis of 536 convalescent healthcare workers reveals that SARS-CoV-2-specific and virus-neutralizing antibody levels are elevated in individuals that experience severe disease. The severity-associated increase in SARS-CoV-2-specific antibody is dominated by immunoglobulin G (IgG), with an IgG subclass ratio skewed toward elevated receptor binding domain (RBD)- and S1-specific IgG3. In addition, individuals that experience severe disease show elevated SARS-CoV-2-specific antibody binding to the inflammatory receptor FcɣRIIIa. Based on these correlational studies, we propose that spike-specific IgG subclass utilization may contribute to COVID-19 disease severity through potent Fc-mediated effector functions. These results may have significant implications for SARS-CoV-2 vaccine design and convalescent plasma therapy.
Assuntos
Anticorpos Antivirais/sangue , COVID-19/sangue , Imunoglobulina G/sangue , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , SARS-CoV-2/imunologia , Índice de Gravidade de DoençaRESUMO
PURPOSE: The purpose of this study was to investigate the stability of reconstituted infliximab solutions and determine whether infliximab is suitable for compounding for potential intravitreal use. METHODS: Infliximab was reconstituted, and the solution was aliquoted and stored refrigerated. On each day of testing, an aliquot was serially diluted to concentrations ranging from 50,000 pg/mL to 69 pg/mL. Each dilution was assayed by microsphere immunoassay daily for 5 days and weekly for a total of 6 weeks. The outcome measure was median fluorescence intensity measured by dual laser flow analysis of fluorochrome-labeled secondary antibodies to infliximab bound to tumor necrosis factor-alpha-coated microspheres. RESULTS: There was an increasing median fluorescence intensity for increasing infliximab concentration in a sigmoidal dose-response curve with a variable slope that was equivalent for each time point. Each respective concentration of infliximab showed nearly equivalent median fluorescence intensity for every time point over the 6-week period. CONCLUSION: The authors found that the immunoreactivity of 2 different concentrations of infliximab stored at 4 degrees C over a 6-week period remained stable. Infliximab is suitable for compounding and could be a cost-effective intravitreal medication for use in clinical practice if further study supports its safety and efficacy.
Assuntos
Anti-Inflamatórios/química , Anticorpos Monoclonais/química , Química Farmacêutica , Anti-Inflamatórios/economia , Anticorpos Monoclonais/economia , Análise Custo-Benefício , Composição de Medicamentos , Custos de Medicamentos , Estabilidade de Medicamentos , Armazenamento de Medicamentos , Infliximab , Fator de Necrose Tumoral alfa/antagonistas & inibidoresRESUMO
BACKGROUND: To determine intraocular levels of bevacizumab in a normal eye 24 hours after the injection of bevacizumab into the vitreous cavity. METHODS: Fluid and tissue samples were analyzed for unbound bevacizumab levels using microsphere immunoassays. RESULTS: After a loading dose of 2.5 mg (0.1 mL) of bevacizumab, levels of unbound drug could be detected in all tissue extracts 24 hours after its injection. At least 60.2% of the injected bevacizumab could be accounted for in this study. Levels of 0.06 mg (2.4%) of bevacizumab were recorded in the choroid. CONCLUSION: At 24 hours, unbound bevacizumab levels can be detected at the level of the choroid. Therefore, bevacizumab can reach the site where choroidal neovascularization develops. This explains its reported therapeutic effect.