Protein Detection Methods in Proteomics Research

Westermeier, R.; Marouga, Rita

doi:10.1007/s10540-005-2845-1

Cited by 126 publications

(77 citation statements)

References 35 publications

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“…Proteins destain to varying extents along with the gel matrix, but with slower kinetics. The exception are some proteins, e.g., collagen, which in the presence of solvent fade more quickly than gel background [15].…”

Section: Introductionmentioning

confidence: 99%

Protein stains for proteomic applications:Which, when, why?

Crawford²,

2006

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This review recollects literature data on sensitivity and dynamic range for the most commonly used colorimetric and fluorescent dyes for general protein staining, and summarizes procedures for the most common PTM-specific detection methods. It also compiles some important points to be considered in imaging and evaluation. In addition to theoretical considerations, examples are provided to illustrate differential staining of specific proteins with different detection methods. This includes a large body of original data on the comparative evaluation of several pre-and post-electrophoresis stains used in parallel on a single specimen, horse serum run in 2-DE (IPG-DALT). A number of proteins/protein spots are found to be over-or under-revealed with some of the staining procedures.

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Section: Introductionmentioning

confidence: 99%

Protein stains for proteomic applications:Which, when, why?

Crawford²,

2006

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“…Still, in addition to the proof of concept of the methodology, it can be assessed that the developed method allows for a limit of detection in the low ng mm À2 range, which is of the same magnitude as those provided by the state of the art protein detection over PVDF membranes [13,48,49]. However the current difference increases with the protein concentration, further experiments will have to be performed to thoroughly characterize the proposed method by for example determining its limit of quantification and linear dynamic range.…”

Section: à2mentioning

confidence: 99%

“…Indeed, detection of proteins, after derivatization [7,8] or by native fluorescence [11], has been reported with detection limits close to 5 pg per band on polyacrylamide gels and between 0.25 and 1 ng mm À2 on polyvinylidene fluoride (PVDF) membranes [12]. Still, this approach is somehow costly as expensive fluorescent dyes and/or fluorescence scanners are required [13]. Electrochemical detection of proteins by scanning electrochemical microscopy (SECM) has been developed in conjunction with techniques like immunodetection [14,15] or metal staining [16][17][18][19].…”

Section: Introductionmentioning

confidence: 99%

Adsorbed protein detection by scanning electrochemical microscopy

Cortés‐Salazar

Busnel

2009

Journal of Electroanalytical Chemistry

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a b s t r a c tA scanning electrochemical microscopy (SECM) protein detection methodology has been developed based on the tagging of free cysteines and other nucleophiles in proteins and peptides by benzoquinone. The tagged proteins are detected by the mediated reduction of benzoquinone with a redox species produced electrochemically at the SECM tip. After careful optimization, a sensitivity in the low ng mm À2 range was reached for bovine serum albumin. One of the major advantages of the present technique is that the selectivity of the protein tagging can be tuned by changing the pH of the reaction media. Depending on the requirements, cysteine selective or general detection can therefore be achieved with a high sensitivity. As a proof of concept, this technique was applied to the detection of protein spots and to the imaging of human fingerprints and further compared to the actual SECM state-of-art approach.

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“…Colloidal Coomassie blue 20 is a background-free stain that can detect as little as 15 ng per 2-DE spot. 21 This stain is generally compatible with mass spectrometry and its use is generally less complex than silver staining. Fluorescent staining with SYPRO Ruby is a recent alternative that is relatively simple to perform, fast, and sensitive.…”

Section: Protein Visualization and Quantitationmentioning

confidence: 99%

Two-dimensional protein electrophoresis: From molecular pathway discovery to biomarker discovery in neurological disorders

Choe

Werner

Lee

2006

NeuroRX

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Summary: Two-dimensional protein electrophoresis (2-DE) has undergone many technical improvements in the past 30 years, resulting in an analytical method that is unparalleled in the resolution of complex protein mixtures and capable of quantifying changes in protein expression from a wide variety of tissues and samples. The technique has been applied in many studies of neurologic disease to identify changes in spot patterns that correlate with disease. The true power of the technique emerges when it is coupled to state-of-the-art methods in mass spectrometry, which enable identification of the protein or proteins contained within a spot of interest on a 2-DE map. Investigators have successfully applied the technique to gain improved understanding of neurologic disease mechanisms in humans and in animal models and to discover biomarkers that are useful in the clinical setting. An important extension to these efforts that has not been realized thus far is the desire to profile changes in protein expression that result from therapy to help relate disease-modifying effects at the molecular level with clinical outcomes. Here we review the major advances in 2-DE methods and discuss specific examples of its application in the study of neurologic diseases.

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Protein Detection Methods in Proteomics Research

Cited by 126 publications

References 35 publications

Protein stains for proteomic applications:Which, when, why?

Protein stains for proteomic applications:Which, when, why?

Adsorbed protein detection by scanning electrochemical microscopy

Two-dimensional protein electrophoresis: From molecular pathway discovery to biomarker discovery in neurological disorders

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