A Flexible System of Enzymatic Analysis

1st Edition - August 28, 1972
Author: Oliver Lowry
Language: English
eBook ISBN:
9 7 8 - 0 - 3 2 3 - 1 5 6 7 1 - 4

A Flexible System of Enzymatic Analysis is a multipurpose manual of laboratory methods that offers a systematic scheme for the analysis of biological materials from the level of… Read more

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A Flexible System of Enzymatic Analysis is a multipurpose manual of laboratory methods that offers a systematic scheme for the analysis of biological materials from the level of the wholeorgan down to the single cell and beyond. It intends to guide the development of methods, the refinement of old ones, and the adaptation in general of methods to almost any scale of sensitivity. This manual is organized into three parts: a general section, one on quantitative histochemistry, and an appendix containing information that may be useful to have at the bench. The general section is comprised of nine chapters that focus on properties of the pyridine nucleotides, kinetics, and glassware, as well as on improvement, modification, adaptation, trouble shooting, and development of methods. This part also describes the preparation of tissues for analysis, the enzymatic cycling methods, and a compendium of 36 metabolite assays. The quantitative histochemistry section is comprised of four chapters that include information on the preparation of frozen-dried material and dissection of samples for analysis; the fishpole balance for weighing samples; and the generalities of analysis with emphasis on the ""oil well technique."" This book will be useful to novices as well as experts who are familiar with other analytical styles.

PrefacePart I General Principles Chapter 1 Pyridine Nucleotides Spectral Absorption and Fluorescence of Reduced Pyridine Nucleotides Limitations of Fluorometry Fluorescence Reference Standards Excitation and Emission Wavelengths and Light Filters Conversion of Pyridine Nucleotides in Alkali into Fluorescent Products Increased Fluorescence from DPNH and TPNH with Strong Alkali Stability and Selective Destruction of Reduced and Oxidized Forms of Pyridine Nucleotides Destruction of DPNH and TPNH in Acid and Stability in Alkali Destruction of DPN+ and TPN+ in Alkali and Stability in Acid Preparation, Standardization, and Storage of Pyridine Nucleotide Solutions Fluorometers Chapter 2 A Kinetic Primer for the Tissue Analyst Nonenzyme Kinetics Enzyme Kinetics Chapter 3 Glassware Constriction Pipettes Tubes Chapter 4 Typical Fluorometric Procedures for Metabolic Assays A One-Step Metabolite Assay with TPN+ A One-Step Metabolite Assay with DPNH A Multistep Metabolite Assay (for Inorganic Phosphate) Chapter 5 Measurement of Enzyme Activities with Pyridine Nucleotides A One-Step Enzyme Assay with TPN+ (Isocitric Dehydrogenase) A One-Step Enzyme Assay with DPNH (Glycero-P Dehydrogenase) A Multistep Enzyme Assay (P-Glucomutase) Chapter 6 Improvement, Modification, Adaptation, Trouble Shooting, and Development of New Methods Simplification and Improvement Adaptation of Pyridine Nucleotide Methods to Increase Sensitivity Development of a New Method Chapter 7 Preparation of Tissues for Analysis Preparation for Enzyme Assays Preparation for Metabolite Analyses Preparation of Tissue Extracts The Use of Methanol-HCl in Preparing Extracts Fluorescence of Tissue Extracts Measurement of Pyridine Nucleotides in Tissues Chapter 8 Enzymatic Cycling TPN Cycle DPN Cycle ATP-ADP Cycle Glutamate-α-ketoglutarate Cycle Other Cycles Double Cycling Chapter 9 A Collection of Metabolite Assays ADP and AMP ATP and P-Creatine CitratePart II Quantitative Histochemistry Introduction Chapter 10 Preparation of Tissues And Sections Freezing Storage of Frozen Tissues Mounting the Tissue for Cutting Sectioning the Tissue Drying of Samples Storage of Dry Sections Chapter 11 Dissection and Histological Control Dissection and Balance Room Removal of Sections from Evacuation Tube Dissection Handling Dissected Samples Identification and Histological Control Sample Carriers Defatting Frozen Dried Samples Sample Volume Chapter 12 The Quartz Fiber Fishpole Balance Measurement of Displacement and Useful Range Choice of Suitable Fiber. Sensitivity as a Function of Length and Diameter of the Fiber Balance Fibers Pans Balance Case Mounting the Fiber Lighting and Viewing Weighing Correction of Weight for Adsorption of Gases and Moisture Calibration of Balances and Testing Linearity Cleaning the Balance Fiber Chapter 13 Histochemical Analysis Introduction Enzyme Stability at High Dilution Addition of Sample and Reagent Oil Well Technique Reagent Wells Pipettes Microscope and Working Stage Addition of Sample and Reagent Special ConsiderationsPart III Appendix Appendix Acid-Base Made Easy Statistical Shortcuts Construction of Dissecting Knives Microtome Knife Sharpening Table of Indicators Preparation and Storage of Common Stock Solutions Normality of Concentrated Acids and Bases Commercial Sources of Equipment and Supplies Abbreviated List of Atomic WeightsReferencesSubject Index