Multiplexed quantification of surface-bound proteins on gold nanoparticles

Hong, Seong-Hyun; Kim, Min ji; Ahn, Joong‐Hoon; Yeo, Woon‐Seok

doi:10.1039/c3ay41037d

Cited by 8 publications

(7 citation statements)

References 20 publications

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“…This result is consistent with a previous report in which 100 BSA molecules (similar in size) were usually observed on individual 40 nm Au nanoparticles. [14] When kinase Src was added to the solution, the SRCtide could be phosphorylated in the presence of ATP, kinase Src, and Mg 2+ , leading to more negative charge of the Au NPs. Black dots in Figure 3 c show the charge change of an individual nanooscillator marked by the white arrow in Figure 3 a,b.…”

Section: Methodsmentioning

confidence: 99%

Real‐Time Monitoring of Phosphorylation Kinetics with Self‐Assembled Nano‐oscillators

et al. 2015

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Phosphorylation is a post-translational modification that is involved in many basic cellular processes and diseases, but is difficult to detect in real time with existing technologies. A label-free detection of phosphorylation is reported in real time with self-assembled nano-oscillators. Each nano-oscillator consists of a gold nanoparticle tethered to a gold surface with a molecular linker. When the nanoparticle is charged, the nano-oscillator can be driven into oscillation with an electric field and detected with a plasmonic imaging approach. The nano-oscillators measure charge change associated with phosphorylation of peptides attached onto a single nanoparticle, allowing us to study the dynamic process of phosphorylation in real time without antibodies down to a few molecules, from which Michaelis and catalytic rate constants are determined.

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Section: Methodsmentioning

confidence: 99%

Real‐Time Monitoring of Phosphorylation Kinetics with Self‐Assembled Nano‐oscillators

et al. 2015

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“…The absolute and multiplexed quantification was performed by analysis of specific peptide fragments which were generated by on bead tryptic digestion of protein‐GNP conjugates using matrix assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS) in the presence of an isotope‐labeled internal standard (Fig. ) . In this study it was found that the surface coverage of the individual proteins is not only related to the nanoparticle size employed as carrier but also the concentration and mixing ratio of the proteins to be immobilized in the reaction mixture.…”

Section: Surface Coverage Determinationmentioning

confidence: 97%

“…Comparison of the intensities between ISs and reference peptide fragments from protein 1 and protein 2 allowed the absolute and multiplexed quantification of surface‐bound proteins. (Reprinted with permission from. © The Royal Society of Chemistry 2013).…”

Section: Surface Coverage Determinationmentioning

confidence: 99%

Functionalized gold nanoparticles for sample preparation: A review

Liu

Lämmerhofer

2019

Electrophoresis

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Sample preparation is a crucial step for the reliable and accurate analysis of both small molecule and biopolymers which often involves processes such as isolation, pre‐concentration, removal of interferences (purification), and pre‐processing (e.g., enzymatic digestion) of targets from a complex matrix. Gold nanoparticle (GNP)‐assisted sample preparation and pre‐concentration has been extensively applied in many analytical procedures in recent years due to the favorable and unique properties of GNPs such as size‐controlled synthesis, large surface‐to‐volume ratio, surface inertness, straightforward surface modification, easy separation requiring minimal manipulation of samples. This review article primarily focuses on applications of GNPs in sample preparation, in particular for bioaffinity capture and biocatalysis. In addition, their most common synthesis, surface modification and characterization methods are briefly summarized. Proper surface modification for GNPs designed in accordance to their target application directly influence their functionalities, e.g., extraction efficiencies, and catalytic efficiencies. Characterization of GNPs after synthesis and modification is worthwhile for monitoring and controlling the fabrication process to ensure proper quality and functionality. Parameters such as morphology, colloidal stability, and physical/chemical properties can be assessed by methods such as surface plasmon resonance, dynamic light scattering, ζ‐potential determinations, transmission electron microscopy, Taylor dispersion analysis, and resonant mass measurement, among others. The accurate determination of the surface coverage appears to be also mandatory for the quality control of functionality of the nanoparticles. Some promising applications of (functionalized) GNPs for bioanalysis and sample preparation are described herein.

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“…The number of molecules immobilized on the surface of a nanoparticle has been measured in several works [10][11][12][13][14][15][16][17][18][19][20][21]. To complete this task, a variety of methods have been proposed: spectroscopic (absorption [11], emission [12], fluorescent [13,14], etc.…”

Section: Introductionmentioning

confidence: 99%

Retention of Activity by Antibodies Immobilized on Gold Nanoparticles of Different Sizes: Fluorometric Method of Determination and Comparative Evaluation

et al. 2021

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Antibody–nanoparticle conjugates are widely used analytical reagents. An informative parameter reflecting the conjugates’ properties is the number of antibodies per nanoparticle that retain their antigen-binding ability. Estimation of this parameter is characterized by a lack of simple, reproducible methods. The proposed method is based on the registration of fluorescence of tryptophan residues contained in proteins and combines sequential measurements of first the immobilized antibody number and then the bound protein antigen number. Requirements for the measurement procedure have been determined to ensure reliable and accurate results. Using the developed technique, preparations of spherical gold nanoparticles obtained by the most common method of citrate reduction of gold salts (the Turkevich–Frens method) and varying in average diameter from 15 to 55 nm have been characterized. It was shown that the number of antibodies (immunoglobulins G) bound by one nanoparticle ranged from 30 to 194 during adsorptive unoriented monolayer immobilization. C-reactive protein was considered as the model antigen. The percentage of antibody valences that retained their antigen-binding properties in the conjugate increased from 17 to 34% with an increase in the diameter of gold nanoparticles. The proposed method and the results of the study provide tools to assess the capabilities of the preparations of gold nanoparticles and their conjugates as well as the expediency of seeking the best techniques for various practical purposes.

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Multiplexed quantification of surface-bound proteins on gold nanoparticles

Cited by 8 publications

References 20 publications

Real‐Time Monitoring of Phosphorylation Kinetics with Self‐Assembled Nano‐oscillators

Real‐Time Monitoring of Phosphorylation Kinetics with Self‐Assembled Nano‐oscillators

Functionalized gold nanoparticles for sample preparation: A review

Retention of Activity by Antibodies Immobilized on Gold Nanoparticles of Different Sizes: Fluorometric Method of Determination and Comparative Evaluation

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