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Optimizing protein solubility for two-dimensional gel electrophoresis analysis of human myocardium. 2003

Brian A Stanley, and Irina Neverova, and Heather A Brown, and Jennifer E Van Eyk
Department of Physiology, Queen's University, Kingston, ON, Canada.

In order to maximize the myocardial proteome observed by two-dimensional gel electrophoresis (2-DE), the effect of (1) either an ionic or different zwitterionic detergents during tissue homogenization and (2) altering the "standard" detergent for isoelectric focusing (3-[(3-cholamidopropyl)dimethylamino]-1-propane sulfonate (CHAPS)) to either the zwitterionic detergent amidosulfobetaine-14 (ASB-14) or N-decyl-N-N'-dimethyl-3-ammonio-1-propane sulfonate (SB3-10) was investigated. Sodium dodecyl sulfate was shown to be a superior detergent for extraction of proteins during homogenization of cardiac tissue compared to the detergents ASB-14, SB3-10 or CHAPS. Additionally, both ASB-14 and SB3-10 exhibited better extraction than CHAPS for distinct regions of two-dimensional gels. In most cases, the best combination of homogenization and focusing conditions did not involve the use of the same detergent. Specifically, it was found that the ability to mix homogenization and focusing conditions can allow one to obtain an optimum balance between the resolution and number of protein spots obtained in 2-DE analysis of cardiac tissue. An excellent initial combination of buffers to utilize for the general examination of cardiac proteins was determined to be initial homogenization in a buffer containing ASB-14 followed by focusing in a buffer containing CHAPS.

UI MeSH Term Description Entries
D007525 Isoelectric Focusing Electrophoresis in which a pH gradient is established in a gel medium and proteins migrate until they reach the site (or focus) at which the pH is equal to their isoelectric point. Electrofocusing,Focusing, Isoelectric
D009124 Muscle Proteins The protein constituents of muscle, the major ones being ACTINS and MYOSINS. More than a dozen accessory proteins exist including TROPONIN; TROPOMYOSIN; and DYSTROPHIN. Muscle Protein,Protein, Muscle,Proteins, Muscle
D009206 Myocardium The muscle tissue of the HEART. It is composed of striated, involuntary muscle cells (MYOCYTES, CARDIAC) connected to form the contractile pump to generate blood flow. Muscle, Cardiac,Muscle, Heart,Cardiac Muscle,Myocardia,Cardiac Muscles,Heart Muscle,Heart Muscles,Muscles, Cardiac,Muscles, Heart
D002021 Buffers A chemical system that functions to control the levels of specific ions in solution. When the level of hydrogen ion in solution is controlled the system is called a pH buffer. Buffer
D003902 Detergents Purifying or cleansing agents, usually salts of long-chain aliphatic bases or acids, that exert cleansing (oil-dissolving) and antimicrobial effects through a surface action that depends on possessing both hydrophilic and hydrophobic properties. Cleansing Agents,Detergent Pods,Laundry Detergent Pods,Laundry Pods,Syndet,Synthetic Detergent,Agent, Cleansing,Agents, Cleansing,Cleansing Agent,Detergent,Detergent Pod,Detergent Pod, Laundry,Detergent Pods, Laundry,Detergent, Synthetic,Detergents, Synthetic,Laundry Detergent Pod,Laundry Pod,Pod, Detergent,Pod, Laundry,Pod, Laundry Detergent,Pods, Detergent,Pods, Laundry,Pods, Laundry Detergent,Synthetic Detergents
D006352 Heart Ventricles The lower right and left chambers of the heart. The right ventricle pumps venous BLOOD into the LUNGS and the left ventricle pumps oxygenated blood into the systemic arterial circulation. Cardiac Ventricle,Cardiac Ventricles,Heart Ventricle,Left Ventricle,Right Ventricle,Left Ventricles,Right Ventricles,Ventricle, Cardiac,Ventricle, Heart,Ventricle, Left,Ventricle, Right,Ventricles, Cardiac,Ventricles, Heart,Ventricles, Left,Ventricles, Right
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D012995 Solubility The ability of a substance to be dissolved, i.e. to form a solution with another substance. (From McGraw-Hill Dictionary of Scientific and Technical Terms, 6th ed) Solubilities
D014020 Tissue Extracts Preparations made from animal tissues or organs (ANIMAL STRUCTURES). They usually contain many components, any one of which may be pharmacologically or physiologically active. Tissue extracts may contain specific, but uncharacterized factors or proteins with specific actions. Extracts, Tissue
D015180 Electrophoresis, Gel, Two-Dimensional Electrophoresis in which a second perpendicular electrophoretic transport is performed on the separate components resulting from the first electrophoresis. This technique is usually performed on polyacrylamide gels. Gel Electrophoresis, Two-Dimensional,Polyacrylamide Gel Electrophoresis, Two-Dimensional,2-D Gel Electrophoresis,2-D Polyacrylamide Gel Electrophoresis,2D Gel Electrophoresis,2D PAGE,2D Polyacrylamide Gel Electrophoresis,Electrophoresis, Gel, 2-D,Electrophoresis, Gel, 2D,Electrophoresis, Gel, Two Dimensional,Polyacrylamide Gel Electrophoresis, 2-D,Polyacrylamide Gel Electrophoresis, 2D,Two Dimensional Gel Electrophoresis,2 D Gel Electrophoresis,2 D Polyacrylamide Gel Electrophoresis,Electrophoresis, 2-D Gel,Electrophoresis, 2D Gel,Electrophoresis, Two-Dimensional Gel,Gel Electrophoresis, 2-D,Gel Electrophoresis, 2D,Gel Electrophoresis, Two Dimensional,PAGE, 2D,Polyacrylamide Gel Electrophoresis, 2 D,Polyacrylamide Gel Electrophoresis, Two Dimensional,Two-Dimensional Gel Electrophoresis

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