Enhanced detectability in proteome studies

Sriyam, Supawadee; Sinchaikul, Supachok; Tantipaiboonwong, Payungsak; Tzao, Ching; Phutrakul, Suree; Chen, Shui Tein

doi:10.1016/j.jchromb.2006.10.065

Cited by 16 publications

(7 citation statements)

References 82 publications

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“…Conventional 2DE permits the separation of protein samples according to their pI and MW [39], and was used herein to initially analyze the physicochemical characteristics of the AF, BF, and CF. The 2DE analyses were performed twice and the representative images were shown in Fig 3A–3D.…”

Section: Resultsmentioning

confidence: 99%

Enhanced Detection of Low-Abundance Human Plasma Proteins by Integrating Polyethylene Glycol Fractionation and Immunoaffinity Depletion

et al. 2016

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The enormous depth complexity of the human plasma proteome poses a significant challenge for current mass spectrometry-based proteomic technologies in terms of detecting low-level proteins in plasma, which is essential for successful biomarker discovery efforts. Typically, a single-step analytical approach cannot reduce this intrinsic complexity. Current simplex immunodepletion techniques offer limited capacity for detecting low-abundance proteins, and integrated strategies are thus desirable. In this respect, we developed an improved strategy for analyzing the human plasma proteome by integrating polyethylene glycol (PEG) fractionation with immunoaffinity depletion. PEG fractionation of plasma proteins is simple, rapid, efficient, and compatible with a downstream immunodepletion step. Compared with immunodepletion alone, our integrated strategy substantially improved the proteome coverage afforded by PEG fractionation. Coupling this new protocol with liquid chromatography-tandem mass spectrometry, 135 proteins with reported normal concentrations below 100 ng/mL were confidently identified as common low-abundance proteins. A side-by-side comparison indicated that our integrated strategy was increased by average 43.0% in the identification rate of low-abundance proteins, relying on an average 65.8% increase of the corresponding unique peptides. Further investigation demonstrated that this combined strategy could effectively alleviate the signal-suppressive effects of the major high-abundance proteins by affinity depletion, especially with moderate-abundance proteins after incorporating PEG fractionation, thereby greatly enhancing the detection of low-abundance proteins. In sum, the newly developed strategy of incorporating PEG fractionation to immunodepletion methods can potentially aid in the discovery of plasma biomarkers of therapeutic and clinical interest.

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Section: Resultsmentioning

confidence: 99%

Enhanced Detection of Low-Abundance Human Plasma Proteins by Integrating Polyethylene Glycol Fractionation and Immunoaffinity Depletion

et al. 2016

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“…9). The quenching rate K SV can be obtained by linear Stern-Volmer plots of F 0 /F against [Q] from (1). As shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Proteins play an important role in various biological processes [1]. Human serum and plasma proteomes are estimated to be composed of more than 10 000 different proteins in a dynamic range of protein concentration, most of which would be present at very low relative abundances [2, 3].…”

Section: Introductionmentioning

confidence: 99%

Hexamethylene‐1,3‐bis(cetyldimethylammonium bromide) protected low‐toxicity ZnSe quantum dots as fluorescent probe for proteins

Zhong

Deng

et al. 2015

Micro & Nano Letters

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Hexamethylene-1,3-bis(cetyldimethylammonium bromide) (G 16-6-16) protected low-toxicity ZnSe quantum dots (QDs) were successfully synthesised and used as fluorescence probe to detect proteins such as bovine serum albumin (BSA). This method is simple, fast, selective and sensitive. Surfactant G 16-6-16 can interact with proteins through hydrophobic effect, thus leading to the fluorescence quenching of G 16-6-16 protected ZnSe QDs nanometre material. The fluorescence intensity of G 16-6-16 protected ZnSe QDs was linearly proportional to BSA over a concentration range from 0 to 80 nM with a correlation coefficient of 0.993. The detection limit was 0.04 nM. This method was successfully applied to determine BSA in urine samples which shows the good application value of this nanometre material.

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“…Therefore, the accurate and sensitive detection of tyrosinase could offer relevant information for a better understanding of different tyrosinaserelated biological processes. The latter represents a demanding task, because tyrosinase, as well as other enzymes, is normally expressed inside cells or tissues in concentration levels that are temporally and spatially dependent [7,8]. In proteomics, a common strategy employed to tackle such situation is based on the extraction, separation and identification of proteins by using protein electrophoresis (e.g.…”

Section: Introductionmentioning

confidence: 99%

Multiple scanning electrochemical microscopy mapping of tyrosinase in micro-contact printed fruit samples on polyvinylidene fluoride membrane

Lin

Cortés‐Salazar

Lesch

et al. 2015

Electrochimica Acta

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Keywords:Scanning electrochemical microscopy tyrosinase enzymatic activity electrochemical immunoassay readout A B S T R A C T Herein, we introduce three orthogonal and compatible methods for detecting tyrosinase, a key factor in fruit browning and skin disorders, with high spatial resolution by means of scanning electrochemical microscopy (SECM). All methods are performed subsequently on the same substrate area providing a wide range of relevant information. The first SECM approach that relies on the mapping of a differential pore oxygen concentration induced by the local hydrophobic changes that the adsorption of tyrosinase generates on a porous polyvinylidene fluoride (PVDF) membrane. The second approach is based on the direct monitoring of the enzymatic activity of tyrosinase by detecting amperometrically the reaction products from the enzymatic conversion of L-3,4-dihydroxyphenylalanine (L-DOPA). Finally, tyrosinase was visualized implementing a tyrosinase sandwich immunoassay readout by SECM. The multiple SECM detection strategies were successfully applied to map unequivocally the tyrosinase enzymatic activity of a micro-contact printed banana sample. Furthermore, differential pulse voltammetry and mass spectrometry analyses were employed to elucidate the nature of the electrochemical response obtained during the tyrosinase enzymatic activity experiments.

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Enhanced detectability in proteome studies

Cited by 16 publications

References 82 publications

Enhanced Detection of Low-Abundance Human Plasma Proteins by Integrating Polyethylene Glycol Fractionation and Immunoaffinity Depletion

Enhanced Detection of Low-Abundance Human Plasma Proteins by Integrating Polyethylene Glycol Fractionation and Immunoaffinity Depletion

Hexamethylene‐1,3‐bis(cetyldimethylammonium bromide) protected low‐toxicity ZnSe quantum dots as fluorescent probe for proteins

Multiple scanning electrochemical microscopy mapping of tyrosinase in micro-contact printed fruit samples on polyvinylidene fluoride membrane

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