Sensitivity and specificity: twin goals of proteomics assays. Can they be combined?

Wilson, Robert

doi:10.1586/epr.13.7

Cited by 65 publications

(62 citation statements)

References 75 publications

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“…where noise σ is the noise floor of measurement and S is the sensitivity of the measurement technique [80]. It is believed that the noise floor in the current generation of biosensors mostly arises from non-specific binding (NSB) processes, for e.g., binding of interfering molecules to the receptors [81].…”

Section: Why Might Label-free Assays Perform Worse Than Labeled Assays?mentioning

confidence: 99%

Performance limitations of label-free sensors in molecular diagnosis using complex samples

Varma

2016

SPIE Proceedings

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Immunoassay formats applicable for clinical or point-of-care diagnostics fall into two broad classes. One which uses labeled secondary antibodies for signal transduction and the other which does not require the use of any labels. Comparison of the limits of detection (LoD) reported by these two sensing approaches over a wide range of detection techniques and target molecules in serum revealed that labeled techniques achieve 2-3 orders of magnitude better LoDs. Further, a vast majority of commercial tests and recent examples of technology translations are based on labeled assay formats. In light of this data, it is argued that extension of traditional labeled approaches and enhancing their functionality may have better clinical impact than the development of newer label-free techniques.

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Section: Why Might Label-free Assays Perform Worse Than Labeled Assays?mentioning

confidence: 99%

Performance limitations of label-free sensors in molecular diagnosis using complex samples

Varma

2016

SPIE Proceedings

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“…It is well-known that many factors can influence results from microarray analysis [98, 99], and the presence of large numbers of antisense transcripts require preparation of directional libraries for RNA-seq analysis [100]. Proteins expressed at low levels are unlikely to be detected using current proteomics technologies [101]. Even with the limitations, combinations of SM screening and genome-wide approaches will provide powerful tools for studying gene function and gene interaction.…”

Section: Figurementioning

confidence: 99%

Chemical genomics for studying parasite gene function and interaction

Li¹,

Yuan²,

Cheng³

et al. 2013

Trends in Parasitology

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With the development of new technologies in genome sequencing, gene expression profiling, genotyping, and high-throughput screening of chemical compound libraries, small molecules are playing increasingly important roles in studying gene expression regulation, gene-gene interaction, and gene function. Here we briefly review and discuss some recent advancements in drug target identification and phenotype characterization using combinations of high-throughput screening of small-molecule libraries and various genome-wide methods such as whole genome sequencing, genome-wide association studies, and genome-wide expressional analysis. These approaches can be used to search for new drugs against parasitic infections, to identify drug targets or drug-resistance genes, and to infer gene function.

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“…5,6 Instead, to achieve high specificity and sensitivity, protein detection assays typically operate via surface-capture of target molecules by affinity reagents such as antibodies or aptamers, which isolate the target protein prior to detection through signal amplification. 7,8 Foremost amongst the techniques that employ this principle are enzyme-linked immunosorbent assays (ELISAs), 9,10 which rely, in their most common implementation, on the surface-capture of target molecules by a dual antibody pair in a "sandwich" complex format, followed by an enzymedriven signal amplification step (Figure 1a). A number of recent approaches have advanced the classical ELISA technique, enabling remarkable improvements in its sensitivity and throughput.…”

Section: Introductionmentioning

confidence: 99%

Direct digital sensing of protein biomarkers in solution

Krainer

Saar

Arter

et al. 2020

Preprint

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Highly sensitive detection of proteins is of central importance to biomolecular analysis and diagnostics. Conventional protein sensing assays, such as ELISAs, remain reliant on surface-immobilization of target molecules and multi-step washing protocols for the removal of unbound affinity reagents. These features constrain parameter space in assay design, resulting in fundamental limitations due to the underlying thermodynamics and kinetics of the immunoprobe–analyte interaction. Here, we present a new experimental paradigm for the quantitation of protein analytes through the implementation of an immunosensor assay that operates fully in solution and realizes rapid removal of excess probe prior to detection without the need of washing steps. Our single-step optofluidic approach, termed digital immunosensor assay (DigitISA), is based on microfluidic electrophoretic separation combined with single-molecule laser-induced fluorescence microscopy and enables calibration-free in-solution protein detection and quantification within seconds. Crucially, the solution-based nature of our assay and the resultant possibility to use arbitrarily high probe concentrations combined with its fast operation timescale enables quantitative binding of analyte molecules regardless of the capture probe affinity, opening up the possibility to use relatively weak-binding affinity reagents such as aptamers. We establish and validate the DigitISA platform by probing a biomolecular biotin–streptavidin binding complex and demonstrate its applicability to biomedical analysis by quantifying IgE–aptamer binding. We further use DigitISA to detect the presence of α-synuclein fibrils, a biomarker for Parkinson’s disease, using a low-affinity aptamer at high probe concentration. Taken together, DigitISA presents a fundamentally new route to surface-free specificity, increased sensitivity, and reduced complexity in state-of-the-art protein detection and biomedical analysis.

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Sensitivity and specificity: twin goals of proteomics assays. Can they be combined?

Cited by 65 publications

References 75 publications

Performance limitations of label-free sensors in molecular diagnosis using complex samples

Performance limitations of label-free sensors in molecular diagnosis using complex samples

Chemical genomics for studying parasite gene function and interaction

Direct digital sensing of protein biomarkers in solution

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