New techniques and updated protocols for the detection and analysis of biomolecules - proteins, glycoproteins and nucleic acids - are presented in the second edition of this successful laboratory manual. Highly sensitive systems which are widely used in molecular biological and biomedical laboratories, such as colorimetric, luminescence, fluorescence measuring using antibody/antigen binding or hybridisation, as well as PCR amplification are described in detail. The clearly structured step-by-step protocols with practical hints and a troubleshooting guide are complemented by chapters on the theoretical background and the application of the techniques. Thus the manual will enable scientists to plan, design and conduct the appropriate procedures which fulfill their particular requirements.

1 General Aspects of Nonradioactive Analysis of Biomolecules

2 Overview of Nonradioactive Labeling Systems
3 Overview on the Digoxigenin:Anti-Digoxigenin (DIG) System
4 Labeling and Detection of Nucleic Acids
5 Labeling and Detection of Proteins and Glycoproteins
6 Labeling and Detection of Nucleic Acids
7 Labeling of Proteins DuringIn VitroTranslation
8 Labeling and Detection of Proteins and Glycoproteins
9 In Vitro Labeling of DNA Probes with 5-BrdU
10 AAF-labeling of DNA and Oligonucleotides
11 Colloidal Gold as a Marker in Molecular Biology: The Use of Ultra-Small Gold Conjugates
12 Direct Peroxidase Labeling of Hybridization Probes and Chemiluminescence Detection
13 The SNAP System
14 Overview on Nonradioactive Detections Systems
15 Overview of Colorimetric, Chemiluminometric, and Fluorimetric Detection Systems
16 Indigo / Tetrazolium Dyes
17 Azo Dyes
18 Chemiluminescent Detection with Horseradish Peroxidase and Luminol
19 Chemiluminescence: Properties of 1,2-Dioxetane Chemiluminescence
20 Electrochemiluminescence: Ruthenium Complexes
21 Bioluminescence: D-Luciferin as Substrate
22 Labeling of Biomolecules with Fluorophores
23 Time-Resolved Fluorescence
24 Overview of Amplification Systems
25 Amplification of Nucleic Acids by Polymerase Chain Reaction: Overview on Basic Principles
26 PCR: Overview on Application Formats in Research and Clinical Diagnosis
27 PCR Amplification for the Generation of DIG-Labeled Probes
28 In Situ PCR Amplification of cDNA
29 SDA Target Amplification
30 In Situ Strand Displacement Amplification
31 Isothermal Amplification of RNA by Transcription-Mediated Amplification (TMA)
32 Ligase Chain Reaction
33 Branched DNA (bDNA) Technology
34 rDNA Amplification: Application of 16S rDNA-Based Methods for Bacterial Identification
35 PRINS: Primed In Situ Labeling and Hybridization in one Step
36 Overview on Amplification Formats
37 Overview on Factors Influencing Nucleic Acid Hybridization
38 Dot, Southern, and Northern Blots
39 PNA Pre-Gel Hybridization - An Alternative to Southern Blotting
40 Multilocus DNA Fingerprinting Using Nonradioactively Labeled Oligonucleotide Probes Specific for Simple Repeat Elements
41 Detection of Proteins and Glycoproteins on Western Blots
42 PNA as Specific Probe for In Situ Hybridization to Metaphase Chromosomes
43 A Highly Sensitive Method for Detecting Peroxidase in In Situ Hybridization or Immunohistochemical Assays
44 Non-Fluorescent Differentiation of Viral and Chromosomal Nucleic Acids in Individual Nuclei
45 Virus Detection in Biopsy Specimens
46 Mapping of Polytene Chromosomes
47 Fluorescence In Situ Hybridization on Banded Chromosomes
48 Spectral Karyotyping: New Tools for Multicolor Chromosome Analysis
49 Chromosome Analysis by Multiplex-FISH (M-FISH)
50 Whole Mount In Situ Hybridization for the Detection of mRNA in Drosophila Embryos
51 Double Labeling of mRNA and Proteins in Drosophila Embryos
52 Detection of DNA/RNA Target/Probe Complexes with DNA/RNA-Specific Antibodies
53 Molecular Beacons: Hybridization Probes for Detection of Nucleic Acids in Homogeneous Solutions
54 Detection of DIG-Labeled Amplicons on Streptavidin-Coated Microtiter Plates
55 A Fluorogenic PCR-Based Assay for the Rapid Detection of Salmonella
56 PCR-Coupled Activation of Intercalating Dyes
57 Simultaneous SDA and Fluorescence Polarization Detection
58 Large-Scale Genomic Sequencing Using Four Color Fluorescent Detection - Principles and Protocols
59 DIG DNA Sequencing with Chemiluminescent or Dye Substrates
60 DNA Sequencing: Chemiluminescent Detection with the 1,2 Dioxane CSPD
61 Direct-Blotting-Electrophoresis (DBE) for DNA Sequencing
62 Sequence Analysis of Nucleic Acids by Mass Spectrometry
63 Mismatch Analysis by PNA Arrays
64 Oligonucleotide Arrays for the Detection of ras Mutations
65 Use of Oligonucleotide Arrays for Gene Expression Monitoring.