Microimmunoassay Using a Protein Chip: Optimizing Conditions for Protein Immobilization

Seong, Seung Yong

doi:10.1128/cdli.9.4.927-930.2002

Cited by 13 publications

(11 citation statements)

References 15 publications

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“…In our hands, they produced three to four times better signal-to-background ratios compared with poly-Llysine slides as well as high signal intensities in the range of a few pg/ml at optimal antibody concentration. In agreement with our results, other studies also reported that epoxysilanized slides possess the highest sensitivity for this type of surface modification (Li and Reichert, 2002;Seong, 2002).…”

Section: One-dimensional Coatingssupporting

confidence: 93%

Solid supports for microarray immunoassays

Kusnezow

Hoheisel

2003

J of Molecular Recognition

266

218

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Stimulated by the achievements of the first phase in genomics and the resulting need of assigning functions to the acquired sequence information, novel formats of immunoassays are being developed for high-throughput multi-analyte studies. In principle, they are similar in nature to the microarray assays already established at the level of nucleic acids. However, the biochemical diversity and the sheer number of proteins are such that an equivalent analysis is much more complex and thus difficult to accomplish. The wide range of protein concentration complicates matters further. Performing microarray immunoassays already represents a challenge at the level of preparing a working chip surface. Arrays have been produced on filter supports, in microtiter plate wells and on glass slides, the last two usually coated with one-, two- or three-dimensionally structured surface modifications. The usefulness and suitability of all these support media for the construction and application of antibody microarrays are reviewed in this manuscript in terms of the different kinds of immunoassay and the various detection procedures. Additionally, the employment of microarrays containing alternative sensor molecules is discussed in this context. The sensitivity of microspot immunoassays predicted by the current analyte theory is not yet a reality, indicating the extent of both the technology's potential and the size of the task still ahead.

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Section: One-dimensional Coatingssupporting

confidence: 93%

Solid supports for microarray immunoassays

Kusnezow

Hoheisel

2003

J of Molecular Recognition

266

218

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“…While this approach may be successful for high abundant or recombinant analytes, a planar substratum cannot attain sufficient surface area per spot for femtomolar detection of analyte proteins in biologically relevant samples. Optimal substrata for protein microarrays must have high binding capacity, high surface area, and intrinsically low background signal (Seong, 2002;Wilson and Nock, 2003). The choice of substratum will dictate the immobilization chemistries employed.…”

Section: Limited Protein Quantitymentioning

confidence: 99%

Protein microarrays: Meeting analytical challenges for clinical applications

et al. 2003

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Protein microarrays, one emerging class of proteomic technologies, have broad applications for discovery and quantitative analysis. A rapidly expanding use of this technology is the acquisition of information about the posttranslational modifications of proteins reflecting the activity state of signal pathways and networks, and is now employed for the analysis of biopsy samples in clinical trial research.

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“…The chips were then placed in a Teflon beaker containing 1% (v/v) (3-glycidyloxypropyl)trimethoxysilane (Sigma-Aldrich, St. Louis, MO) in methanol for a 10-min reaction. While several articles have summarized the surface chemistries for antibody immobilization, [21][22][23][24] the generic protocol involves soaking the chips with silane solution for multiple hours or even overnight to maximize the epoxy density which leaves little hydroxyl group on the SiO 2 surface. The transient 10-min reaction creates a moderate silane surface density, which not only provides covalent linkage but also creates proper surface hydrophilicity for antibody spot diameters being controlled within 90 to 120 lm.…”

Section: Methodsmentioning

confidence: 99%

Multiplexed immunosensing and kinetics monitoring in nanofluidic devices with highly enhanced target capture efficiency

et al. 2016

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Nanofluidic devices promise high reaction efficiency and fast kinetic responses due to the spatial constriction of transported biomolecules with confined molecular diffusion. However, parallel detection of multiple biomolecules, particularly proteins, in highly confined space remains challenging. This study integrates extended nanofluidics with embedded protein microarray to achieve multiplexed real-time biosensing and kinetics monitoring. Implementation of embedded standard-sized antibody microarray is attained by epoxy-silane surface modification and a room-temperature low-aspect-ratio bonding technique. An effective sample transport is achieved by electrokinetic pumping via electroosmotic flow. Through the nanoslit-based spatial confinement, the antigen-antibody binding reaction is enhanced with $100% efficiency and may be directly observed with fluorescence microscopy without the requirement of intermediate washing steps. The image-based data provide numerous spatially distributed reaction kinetic curves and are collectively modeled using a simple one-dimensional convection-reaction model. This study represents an integrated nanofluidic solution for real-time multiplexed immunosensing and kinetics monitoring, starting from device fabrication, protein immobilization, device bonding, sample transport, to data analysis at P eclet number less than 1. Published by AIP Publishing.

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Microimmunoassay Using a Protein Chip: Optimizing Conditions for Protein Immobilization

Cited by 13 publications

References 15 publications

Solid supports for microarray immunoassays

Solid supports for microarray immunoassays

Protein microarrays: Meeting analytical challenges for clinical applications

Multiplexed immunosensing and kinetics monitoring in nanofluidic devices with highly enhanced target capture efficiency

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