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Rapid, highly sensitive, and high-throughput detection of biomarkers at low concentrations is invaluable for early diagnosis of various diseases. In many highly sensitive immunoassays, magnetic beads are used to capture fluorescently labeled target molecules. The target molecules are then quantified by detecting the fluorescent signal from individual beads, which is time consuming and requires a complicated and expensive detection system. Here, we demonstrate a high-throughput optical modulation biosensing (ht-OMB) system, which uses a small permanent magnet to aggregate the beads into a small detection volume and eliminates background noise by steering a laser beam in and out of the cluster of beads. Shortening the aggregation, acquisition, and well-to-well scanning transition times enables reading a 96-well plate within 10 min. Using the ht-OMB system to detect human Interleukin-8, we demonstrated a limit of detection of 0.14 ng/L and a 4-log dynamic range. Testing 94 RNA extracts from 36 confirmed RT-qPCR SARS-CoV-2-positive patients (Ct≤40) and 58 confirmed RT-qPCR SARS-CoV-2-negative individuals resulted in 100% sensitivity and 100% specificity.
Assuntos
COVID-19 , SARS-CoV-2 , Biomarcadores , Humanos , Imunoensaio/métodos , RNA Viral/análise , Sensibilidade e EspecificidadeRESUMO
In many sensitive assays, target molecules are tagged using fluorescently labeled probes and captured using magnetic beads. Here, we introduce an optical modulation biosensing (OMB) system, which aggregates the beads into a small detection area and separates the signal from the background noise by manipulating the laser beam in and out of the cluster of beads. Using the OMB system to detect human interleukin-8, we demonstrated a limit of detection of 0.02 ng/L and a 4-log dynamic range. Using Zika-positive and healthy individuals' serum samples, we show that the OMB-based Zika IgG serological assay has 96% sensitivity and 100% specificity.
RESUMO
Sensitive serological assays are needed to provide valuable information about acute and past viral infections. For example, detection of anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) IgG antibodies could serve as the basis for an "immunity passport" that would enable individuals to travel internationally. Here, utilizing a novel Magnetic Modulation Biosensing (MMB) system and the receptor-binding domain of the SARS-CoV-2 spike protein, we demonstrate a highly sensitive and specific anti-SARS-CoV-2 IgG serological assay. Using anti-SARS-CoV-2 IgG antibodies, RT-qPCR SARS-CoV-2-positive and healthy patients' samples, and vaccinees' samples, we compare the MMB-based SARS-CoV-2 IgG assay's analytical and clinical sensitivities to those of the enzyme-linked immunosorbent assay (ELISA). Compared with ELISA, the MMB-based assay has an ~6-fold lower limit of detection (129 ng/L vs. 817 ng/L), and it detects an increase in the IgG concentration much earlier after vaccination. Using 85 RT-qPCR SARS-CoV-2-positive samples and 79 -negative samples, the MMB-based assay demonstrated similar clinical specificity (98% vs. 99%) and sensitivity (93% vs. 92%) to the ELISA test, but with a much faster turnaround time (45 min vs. 245 min). The high analytical and clinical sensitivity, short turnaround time, and simplicity of the MMB-based assay makes it a preferred method for antibody detection.
Assuntos
Anticorpos Antivirais/análise , Técnicas Biossensoriais , COVID-19 , Imunoglobulina G/análise , Testes Sorológicos , COVID-19/diagnóstico , COVID-19/imunologia , Ensaio de Imunoadsorção Enzimática , Humanos , Fenômenos Magnéticos , SARS-CoV-2/imunologia , Sensibilidade e Especificidade , Glicoproteína da Espícula de CoronavírusRESUMO
BACKGROUND: Cheyne-Stokes breathing (CSB) has been associated with heart failure (HF) patients for many years; however, its true prevalence and its prognostic implications are still obscure. HYPOTHESIS: The goal of this study was to investigate the prevalence and the possible prognostic implications of nocturnal CSB in advanced heart failure patients. METHODS: We performed single night full polysomonography ambulatory sleep studies in 71 HF patients. We analyzed the patients' sleep studies, clinical and laboratory data, and 6 month mortality. RESULTS: A total of 71 chronic systolic HF patients were analyzed, 60 males, 11 females, age 65 +/- 13 years. Mean left ventricular ejection fraction was 27% +/- 11%. Short episodes of CSB (at least 3 min duration) were present in all patients, and mean CSB duration was 1 hour. CSB duration was associated significantly with both high serum levels of N-terminal prohormone brain natriuretic peptide (NT-proBNP) as well as with 6 month mortality. Log CSB time had a significant correlation with log NT-proBNP (r = 0.5, P<.0001). Based on median CSB duration, the Kaplan-Meier survival curve analysis showed significant association with 6 month mortality (P = .03). CONCLUSIONS: CSB prevalence in advanced HF patients is higher than previously reported and is associated with increased serum levels of NT-proBNP and higher 6 month mortality.
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Respiração de Cheyne-Stokes/epidemiologia , Insuficiência Cardíaca Sistólica/epidemiologia , Idoso , Respiração de Cheyne-Stokes/fisiopatologia , Progressão da Doença , Eletrocardiografia Ambulatorial , Feminino , Indicadores Básicos de Saúde , Insuficiência Cardíaca Sistólica/mortalidade , Humanos , Israel/epidemiologia , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Monitorização Ambulatorial , Peptídeo Natriurético Encefálico , Fragmentos de Peptídeos , Polissonografia , Estudos Prospectivos , Fatores de Risco , Índice de Gravidade de Doença , Síndromes da Apneia do Sono , Estatística como Assunto , Volume Sistólico , Função Ventricular EsquerdaRESUMO
Protein-protein interfaces are regions between 2 polypeptide chains that are not covalently connected. Here, we have created a nonredundant interface data set generated from all 2-chain interfaces in the Protein Data Bank. This data set is unique, since it contains clusters of interfaces with similar shapes and spatial organization of chemical functional groups. The data set allows statistical investigation of similar interfaces, as well as the identification and analysis of the chemical forces that account for the protein-protein associations. Toward this goal, we have developed I2I-SiteEngine (Interface-to-Interface SiteEngine) [Data set available at http://bioinfo3d.cs.tau.ac.il/Interfaces; Web server: http://bioinfo3d.cs.tau.ac.il/I2I-SiteEngine]. The algorithm recognizes similarities between protein-protein binding surfaces. I2I-SiteEngine is independent of the sequence or the fold of the proteins that comprise the interfaces. In addition to geometry, the method takes into account both the backbone and the side-chain physicochemical properties of the interacting atom groups. Its high efficiency makes it suitable for large-scale database searches and classifications. Below, we briefly describe the I2I-SiteEngine method. We focus on the classification process and the obtained nonredundant protein-protein interface data set. In particular, we analyze the biological significance of the clusters and present examples which illustrate that given constellations of chemical groups in protein-protein binding sites may be preferred, and are observed in proteins with different structures and different functions. We expect that these would yield further information regarding the forces stabilizing protein-protein interactions.