Biochemical Techniques

, by Robyt, John F.; White, Bernard J.

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ISBN: 9780881335569 | 0881335568
Cover: Hardcover
Copyright: 1/1/1990

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Supplemental Materials

Analyzing and Reporting Experimental Data

(20)

Recording and Handling Experimental Data

(4)

Significant Figures

(2)

Scientific Notation

(1)

Units

(2)

Analysis of Experimental Data

(4)

Error Analysis and the Estimation of Precision

(3)

Errors and the Rejection of Data

(1)

Calculating the Uncertainty of the Result of Mathematical Operations

(1)

The Relation Between Precision and the Number of Determinations

(1)

Tables and Graphs

(8)

Tables

(1)

The Use of Powers of 10 in Tables and Figures

(1)

Rules for Making Graphs

(2)

Linearization

13 (

(1)

Numerical Curve Fitting: The Method of Least Squares

(2)

Weighting Factors in Linear Least Squares

(1)

Calculating the Area Under a Curve

(1)

Controls and Blanks

(1)

General Rules for Nomenclature

(1)

The Laboratory or Research Report

(1)

Literature Cited

(1)

References for Further Study

(2)

Preparation and Properties of Solutions

(19)

Methods of Measurement

(1)

Volumetric Glassware

(5)

Use of All Volumetric Glassware

(1)

Cleaning

(1)

Reading the Meniscus

(1)

Calibration

(1)

The Analytical Balance

(2)

Expression of Concentration and Dilution

(1)

Making Dilutions

(2)

Standard Solutions

(1)

pH and Buffers

(5)

(2)

Buffers

(1)

Buffer Capacity and Ionic Strength

(2)

pH Measurement

(2)

pH Meters

(1)

Making the pH Measurement

(1)

Literature Cited

(1)

References for Further Study

(1)

Spectroscopic Methods

(33)

Electromagnetic Radiation and Spectra

(4)

Quantitative Aspects of Light Absorption

(7)

The Lambert-Beer Law

(1)

Definition of Extinction Coefficients

(1)

Determining an Ultraviolet or Visible Spectrum of a Substance and the Wavelength of Maximum Absorption

(1)

Determining an Extinction Coefficient and concentration

(3)

Determining the Concentration of Two Substances in a Mixture

(1)

Difference Spectroscopy

(1)

Ultraviolet and Visible Spectrometry

(1)

Instrumentation

(1)

Applications of Ultraviolet and Visible Spectroscopy

(1)

Fluorescence Photometry

(4)

Fluorescence

(1)

Instrumentation

(3)

Infrared Spectroscopy

(4)

Infrared Spectrophotometry

(2)

Infrared Spectrophotometers

(1)

Sample Handling for Infrared Spectroscopy

(1)

Applications of Infrared Spectroscopy

(1)

Nuclear Magnetic Resonance Spectrometry

(11)

Proton NMR Spectrometry

(3)

Carbon-13 NMR Spectrometry

(1)

NMR Spectra of Other Nuclei

(3)

Uses of NMR in Biochemistry

(1)

Literature Cited

(1)

References for Further Study

(1)

Chromatographic Techniques

(1)

PART I. TYPES OF CHROMATOGRAPHY AND CHROMATOGRAPHIC TECHNIQUES

(31)

Adsorption Chromatography

(5)

Ion-Exchange Chromatography

(2)

Liquid-Liquid Partition Chromatography

(1)

Paper Chromatography

(5)

Thin-Layer Chromatography

(1)

Gel Permeation Chromatography

(7)

Preparation of Gels and Columns

(1)

Definitions Used in Gel Permeation Chromatography

(1)

Affinity Chromatography

(4)

The Inert Support and the Ligand

(1)

Attachment of the Ligand to the Solid Support

(2)

Hydrophobic Chromatography

(1)

Gas-Liquid Chromatography

(2)

High Performance Liquid Chromatography

101

(3)

PART II. SPECIFIC APPLICATIONS OF CHROMATOGRAPHY

104

(280)

Carbohydrates

104

(7)

Adsorption Chromatography of Carbohydrates

104

(1)

Ion-Exchange Chromatography of Carbohydrates

105

(1)

Paper Chromatography of Carbohydrates

106

(1)

Thin-Layer Chromatography of Carbohydrates

107

(2)

Gel Permeation Chromatography of Carbohydrates

109

(1)

Affinity Chromatography of Carbohydrates

109

(1)

Gas-Liquid Chromatography of Carbohydrates

109

(2)

High Performance Liquid Chromatography of Carbohydrates

111

(1)

Amino Acids

111

(4)

Ion-Exchange Chromatography of Amino Acids

111

(1)

Thin-Layer Chromatography of Amino Acids

112

(2)

High Performance Liquid Chromatography of Amino Acids

114

(1)

Proteins

115

(3)

Adsorption Chromatography of Proteins

115

(1)

Ion-Exchange Chromatography of Proteins

116

(1)

Gel Permeation Chromatography of Proteins

116

(2)

High Performance Liquid Chromatography of Proteins

118

(1)

Nucleotides and Nucleic Acids

118

(3)

Ion-Exchange Chromatography of Nucleic Acids

118

(1)

Gel Permeation of Nucleic Acids

119

(1)

Affinity Chromatography of Nucleic Acids

119

(1)

High Performance Liquid Chromatography of Nucleic Acids

120

(1)

Lipids

121

(3)

Adsorption Chromatography of Lipids

121

(1)

Ion-Exchange Chromatography of Lipids

121

(1)

Paper Chromatography of Lipids

121

(1)

Thin-Layer Chromatography of Lipids

121

(2)

Gas-Liquid Chromatography of Lipids

123

(1)

High Performance Liquid Chromatography of Lipids

123

(1)

Literature Cited

124

(4)

References for Further Study

128

(1)

Electrophoretic Techniques

129

(29)

Theory

129

(1)

Types of Electrophoresis

130

(20)

Paper and Cellulose Acetate Electrophoresis

130

(3)

Thin-Layer Electrophoresis

133

(1)

Gel Electrophoresis

133

(13)

Immunoelectrophoresis

146

(2)

Isoelectric Focusing

148

(2)

Specific Applications

150

(6)

Proteins and Peptides

150

(1)

Nucleic Acids and Oligonucleotides

151

(5)

Lipoprotein Separation

156

(1)

Literature Cited

156

(1)

References for Further Study

157

(1)

Theory, Measurement, and Use of Radioisotopes

158

(55)

Nature of Atoms, Isotopes, and Radioisotopes

158

(1)

Types of Radioactive Decay

159

(3)

Decay by Negative-Beta (Electron) Emission

159

(1)

Decay by Positive-Beta (Positron) Emission

160

(1)

Decay by Electron Capture

160

(1)

Decay by &gamma:-Radiation

161

(1)

Decay by &alph;-Particle Emission

161

(1)

Occurrence and Artificial Production of Radioactive Isotopes

162

(1)

Properties of Radioactive Emissions

163

(8)

Energy of β-Particle Emissions

163

(1)

Interactions of β-Particles with Their Environment

164

(1)

Gamma Radiation and Its Interaction with Matter

165

(1)

Kinetics of Radioactive Decay

166

(5)

Units Used in Radioactive Measurements

171

(1)

The Electron Volt

171

(1)

The Curie: The Unit of Radioactive Disintegration

171

(1)

Special Techniques and Safety in Handling Radioisotopes

172

(1)

Statistics of Counting Radioisotopes

173

(3)

Measurement of Radioactivity by Gas Ionization

176

(4)

Measurement of Radioactivity by Scintillation Counting

180

(10)

Mechanism of Liquid Scintillation Counting

180

(1)

Pulse Height Analysis and the Beta Energy Spectrum

181

(1)

Determining the Activity of Each Isotope in a Dual-Labeled Mixture

182

(2)

Cocktail Composition and Sample Preparation

184

(2)

Quenching and Quench Correction

186

(4)

Solid Scintillation Counting of γ-Radiation

190

(1)

Detection by Autoradiography

191

(2)

Methods of Labeling Biochemical Compounds

193

(4)

Types of Radiochemical Labeling

197

(2)

Applications in the Use of Radioisotopes

199

(10)

The Quantitative Determination of Compounds by Use of Radioisotopes

200

(1)

The Quantitative Determination of a Compound by Isotope Dilution Analysis

201

(3)

The Determination of the Mole Fractions of Several Compounds Formed from One Labeled Precursor

204

(1)

Pulse and Chase Techniques with Radioisotopes

205

(1)

The Use of Radioactive Tracers in the Study of Enzyme Reaction Mechanisms and Metabolic Pathways

206

(1)

Radioimmunoassay

207

(2)

Designing a Radioisotope Experiment

209

(3)

Literature Cited

212

(1)

References for Further Study

212

(1)

Qualitative and Quantitative Methods for Determining Biological Molecules

213

(40)

Carbohydrates

213

(14)

Qualitative Tests for Different Types of Carbohydrates

213

(4)

The Quantitative Determination of Carbohydrates

217

(10)

Amino Acids, Peptides, and Proteins

227

(12)

Qualitative Tests for Amino Acids and Proteins

227

(2)

The Quantitative Determination of Amino Acids

229

(2)

The Quantitative Determination of Proteins

231

(8)

Nucleotides and Nucleic Acids

239

(4)

The Qualitative Determination of RNA and DNA

240

(1)

The Quantitative Determination of Nucleotides and Nucleic Acids

241

(2)

Lipids and Steroids

243

(4)

Qualitative Tests for Lipids on Thin-Layer Chromatograms

243

(2)

Qualitative Tests for Steroids

245

(1)

The Quantitative Determination of Lipids

246

(1)

Miscellaneous

247

(3)

The Determination of Inorganic Phosphate: Modified Fiske--Subbarow Method

247

(1)

Specific Binding Assays

247

(3)

Literature Cited

250

(2)

References for Further Study

252

(1)

Biological Preparations

253

(38)

The Starting Material

253

(1)

Cell Lysis and Extraction

254

(2)

Special Techniques in Preparing Biological Materials

256

(9)

Centrifugation

256

(3)

Density Gradient Centrifugation

259

(1)

Filtration

259

(2)

Concentration of Solutions

261

(2)

Dialysis

263

(1)

Growth of Bacteria for the Production of Biochemical Compounds

264

(1)

Subcellular Fractionation of Cell Components and Organelles

265

(2)

Preparation of Liver Mitochondria

265

(1)

Preparation of Chlorophasts

266

(1)

Isolation of Liver Nuclei

266

(1)

Preparation of Escherichia Coli Ribosomes

267

(1)

Purification of Proteins

267

(8)

Separations Based on Solubility

268

(2)

Separation by Chromatography and Electrophoresis

270

(1)

Crystallization of Proteins

270

(1)

Isolation and Purification of Specific Proteins

271

(4)

Preparation of Carbohydrates

275

(3)

General Separation and Purification Methodology

275

(1)

Specific Methods for Preparing Different Carbohydrates

275

(3)

Preparation of Nucleic Acids

278

(6)

General Separation and Purification Methodology

278

(2)

Specific Methods for Preparing Different Nucleic Acids

280

(4)

Preparation of Lipids

284

(5)

General Separation and Purification Methodology

284

(1)

General Extraction and Purification Procedures

284

(1)

Specific Methods for Preparing Lipids

285

(4)

Literature Cited

289

(1)

References for Further Study

290

(1)

Enzymology

291

(30)

Kinetics and Theory of Enzyme Action

292

(3)

Determining the Initial Velocity vi

295

(1)

Enzyme Reactions as a Function of pH

296

(1)

Enzyme Reactions as a Function of Temperature

297

(1)

Determining the Activity of an Enzyme

298

(6)

Determining the Michaelis Constant Km, the Maximum Velocity Vm and the Turnover Number k2 of an Enzyme

304

(2)

Inhibition of Enzymes

306

(10)

Competitive Inhibition

307

(3)

Noncompetitive Inhibition

310

(1)

Mixed Inhibition

310

(3)

Uncompetitive Inhibition

313

(3)

Allosteric Enzymes That Do Not Follow Michaelis-Menton Kinetics

316

(2)

Specificity of Enzymes

318

(1)

Literature Cited

318

(2)

References for Further Study

320

(1)

Structural Analysis of Biological Molecules

321

(63)

Determining the Structure of Oligosaccharides and Polysaccharides

322

(16)

Determining the Monosaccharide Composition

323

(1)

Determining the Structure of Oligosaccharides

324

(10)

Determining the Structure of Polysaccharides

334

(4)

Determining the Primary Structure of Peptides and Proteins

338

(14)

The Formation and Separation of Polypeptide Chains

339

(1)

Establishing the Purity of the Protein

340

(1)

Determining the Amino Acid Composition

340

(1)

End-Group Analysis

341

(2)

Specific Methods of Peptide Bond Cleavage

343

(3)

The Separation of Peptides

346

(1)

The Sequencing of Peptides

347

(5)

Deducing the Total Sequence from Overlapping Peptides

352

(1)

Determining the Structure of Nucleic Acids

352

(20)

The Isolation, Purification, and Cloning of DNA

353

(2)

The Enzymatic Cleavage of DNA and Restriction Mapping

355

(3)

The Establishment of Purity and the Separation of DNA Strands by Electrophoresis

358

(1)

Sequencing Strategies

358

(14)

Deducing the Complete Sequence for Large Nucleic Acids

372

(1)

Determining the Structure of Lipids

372

(9)

Specific Cleavage of Lipids into Component Parts

374

(1)

Determination of Lipid Components

375

(3)

The Characterization of Phosphatidyl Choline from Egg Yolk---An Example of Lipid Analysis

378

(3)

Literature Cited

381

(3)

Appendix A: Literature Sources for Biochemical Analyses, Methods, and Preparations

384

(3)

Appendix B: Amino Acids, Abbreviations, and DNA Triplet Codes

387

(1)

Appendix C: Some Commercially Available Restriction Endonucleases and Their DNA Sequence Specificities

388

(2)

Appendix D: Table for Preparing Solutions of Different Concentrations of Ammonium Sulfate

390

(3)

Index

393

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Biochemical Techniques

Table of Contents

Supplemental Materials