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Detection of Blotted Proteins

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Cover of 'Detection of Blotted Proteins'

Table of Contents

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    Book Overview
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    Chapter 1 Western Blotting: Origin and Ascent of the Species.
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    Chapter 2 Methods to Concentrate Proteins for Protein Isolation, Proteomic, and Peptidomic Evaluation.
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    Chapter 3 Measuring Protein Concentration on Nitrocellulose and After the Electrophoretic Transfer of Protein to Nitrocellulose
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    Chapter 4 Detection of Blotted Proteins: Not All Blockers Are Created Equal
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    Chapter 5 Protein Stains to Detect Antigen on Membranes
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    Chapter 6 Fluorescent Labeling of Proteins and Its Application to SDS-PAGE and Western Blotting
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    Chapter 7 Rapid, Antibody-Free Detection of Recombinant Proteins on Blots Using Enzyme Fragment Complementation
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    Chapter 8 Use of Nonradioactive Detection Method for North- and South-Western Blot
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    Chapter 9 Immunoblotting Using Radiolabeled Reagents for Detection
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    Chapter 10 Immunoblotting of Antigens: Whole, Strip, and New-Line Nitrocellulose Membrane Immunoblotting Using the Chemiluminescence Technique
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    Chapter 11 Detection of Protein Carbonyls by Means of Biotin Hydrazide–Streptavidin Affinity Methods
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    Chapter 12 Direct Immunodetection of Antigens Within the Precast Polyacrylamide Gel
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    Chapter 13 Quantitative Analysis of Signal Transduction with In-Cell Western Immunofluorescence Assays
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    Chapter 14 Ultrasensitive Protein Detection on Dot Blots and Western Blots with Semiconducting Polymer Dots
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    Chapter 15 Co-detection of Target and Total Protein by CyDye Labeling and Fluorescent ECL Plex Immunoblotting in a Standard Proteomics Workflow.
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    Chapter 16 Using Biotinylated Proteins to Demonstrate Immunodetection of Antigens via Western Blotting, Dot Blots, and Immunohistochemistry
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    Chapter 17 Calcium Binding by Ro 60 Multiple Antigenic Peptides on PVDF Membrane
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    Chapter 18 Sequential Use of Immunoblots for Characterization of Autoantibody Specificities
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    Chapter 19 Nanogold Immunodetection Detection Systems for the Identification of Autoantigens by Western Blotting
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    Chapter 20 Application of Intermittent Microwave Irradiation to Western Blot Analysis
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    Chapter 21 Visualization of Unstained Protein Bands on PVDF
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    Chapter 22 Multiplexed Fluorescent Immunodetection Using Low Autofluorescence Immobilon ® -FL Membrane
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    Chapter 23 Cold Microwave-Enabled Protein Detection and Quantification
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    Chapter 24 TLC-Blot (Far-Eastern Blot) and Its Application to Functional Lipidomics
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    Chapter 25 Analysis of Electroblotted Proteins by Mass Spectrometry.
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    Chapter 26 On-Membrane Renaturation of Recombinant Ro60 Autoantigen by Calcium Ions
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    Chapter 27 Phosphoprotein Detection on Protein Electroblot Using a Phosphate-Specific Fluorophore
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    Chapter 28 Purification of Tryptic Digests on Polyvinylidene Difluoride Membrane
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    Chapter 29 Detection of Blotted Proteins on Nitrocellulose/PVDF Membranes by Alta
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    Chapter 30 Nonstripping “Rainbow” and Multiple Antigen Detection (MAD) Western Blotting
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    Chapter 31 Supported Molecular Matrix Electrophoresis
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    Chapter 32 Parafilm-M(®), An Available Cost-Effective Alternative for Immuno-blot Pouches.
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    Chapter 33 Succinylation-Alcian Blue Staining of Mucins on Polyvinylidene Difluoride Membranes
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    Chapter 34 Comparison of Chemiluminescence vs. Infrared Techniques for Detection of Fetuin-A in Saliva
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    Chapter 35 A Novel Methodology for Stripping and Reprobing of Western Blots Originally Developed with Colorimetric Substrate TMB
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    Chapter 36 Other Notable Methods of Membrane Protein Detection: A Brief Review
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    Chapter 37 Nitrocellulose Membrane: The New Canvas
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    Chapter 38 Invisible Ink Marking in ECL Membrane Assays.
Attention for Chapter 34: Comparison of Chemiluminescence vs. Infrared Techniques for Detection of Fetuin-A in Saliva
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Chapter title
Comparison of Chemiluminescence vs. Infrared Techniques for Detection of Fetuin-A in Saliva
Chapter number 34
Book title
Detection of Blotted Proteins
Published in
Methods in molecular biology, January 2015
DOI 10.1007/978-1-4939-2718-0_34
Pubmed ID
Book ISBNs
978-1-4939-2717-3, 978-1-4939-2718-0
Authors

Suresh T. Mathews, Emily Graff, Robert L. Judd, Vishal Kothari

Abstract

The western blotting technique for transfer and detection of proteins, named following the discovery of southern and northern blotting for DNA- and RNA-blotting, respectively, has traditionally relied on the use of X-ray films to capture chemiluminescence. Recent advancements use super-cooled charge coupled devices (CCD) cameras to capture both chemiluminescence and fluorescence images, which exhibit a greater dynamic range compared to traditional X-ray film. Chemiluminescence detected by a CCD camera records photons and displays an image based on the amount of light generated as a result of a dynamic chemical reaction. Fluorescent detection with a CCD camera, on the other hand, is measured in a static state. Despite this advantage, researchers continue to widely use chemiluminescent detection methods due to the generally poor performance of fluorophores in the visible spectrum. Infrared imaging systems offer a solution to the dynamic reactions of chemiluminescence and the poor performance of fluorophores detected in the visible spectrum, by imaging fluorophores in the infrared spectrum. Infrared imaging is static, has a wide linear range, high sensitivity, and reduced autofluorescence and light scatter. A distinct advantage of infrared imaging is the ability to detect two target proteins simultaneously on the same blot which increases accuracy of quantification and comparison, while minimizing the need for stripping and reprobing. Here, we compare the methodology for chemiluminescent (UVP BioChemi) and infrared (UVP Odyssey) detection of salivary total and phosphorylated fetuin-A, a multifunctional protein associated with cardio-metabolic risk, and discuss the advantages and disadvantages of these methodologies.

Mendeley readers

The data shown below were compiled from readership statistics for 9 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
France 1 11%
Unknown 8 89%

Demographic breakdown

Readers by professional status Count As %
Student > Ph. D. Student 2 22%
Student > Master 2 22%
Student > Bachelor 1 11%
Professor 1 11%
Researcher 1 11%
Other 1 11%
Unknown 1 11%
Readers by discipline Count As %
Medicine and Dentistry 3 33%
Biochemistry, Genetics and Molecular Biology 3 33%
Psychology 1 11%
Pharmacology, Toxicology and Pharmaceutical Science 1 11%
Engineering 1 11%
Other 0 0%