Techniques in Quantification and Localization of Gene Expression - Rilegato

Rilegato

ISBN 10: 0817640347 ISBN 13: 9780817640347

Casa editrice: Springer Basel AG, 2000

Riassunto

The molecular basis of disease is the driving force in biomedical research. Gene detection, quantification, and localization are required to determine disease pathogenesis. In as much as DNA is the genetic potential in a particular organism, RNA and protein expression are the functional result of this genetic programme. This book describes methods to quantify and localize gene expression, with a particular emphasis on the sensitive detection of low abundance gene expression. Sensitive detection of gene expression has been hindered by poor recovery or preservation of target sequences. The text describes optimized methods to ensure full recovery and preservation of targets from specimen acquisition through detection of specific targets and final analysis. This text also focuses upon in situ analysis of cells and tissues

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Contenuti

Basic Methods.- 1. Basic Flow Cytometry Instrumentation.- Instrumentation.- Overview.- Delivery of Sample.- Light Source and Signal Detection.- Data Analysis.- Gating.- Color Compensation.- Conclusion.- 2. Fixation for In Situ Molecular Analysis.- Types of Fixatives.- Cells.- Tissue.- 3. A Method for Reduction of Green Wavelength Autofluorescence Emission in Cellular Preparations.- Protocols―Flow Cytometry.- Trypan Blue with Surface Staining.- Surface Stain Plus In Situ Hybridization.- Surface Label Plus In Situ PCR or RT-PCR.- 4. Gene Quantification: Choosing the Target.- Fresh Cells and Tissue.- Quantification of Gene Expression Prior to In Situ Analysis.- 5. Detection and Quantification of Cytokine-Producing Cells by Immunostaining.- Materials.- Methods.- Procedure 1: Cytokine Detection of Individual Cells Smeared on Slides Using Immunostaining.- Quantification of Individual Cytokine-Producing Cells by Automated Computerized Image Analysis (ACIA).- Assessment of Cell Size, Intensity of Signal, and Enumeration of Individual Cytokine-Producing Cells.- Procedure 2: Immunofluorescent Staining of Cytokine-Producing Cells in Suspension.- Procedure 3. Cytokine Detection in Tissue Using Immunohistochemistry.- Troubleshooting.- 6. In Situ Hybridization Using the bDNA Technology.- Materials and Methods.- Reagents.- Preparation of Cytospin Slides.- Pretreatment of Slides Prior to Hybridization.- Hybridization and Signal Development.- Results and Discussion.- Conclusions.- Acknowledgments.- Specific Applications.- 7. Kaposi’s Sarcoma―Associated Human Herpes Virus Eight: Localization of Viral Gene Expression and Viral Latency.- Materials and Methods.- Clinical Samples.- Solution-Based PCR and Serologic Assays.- In Situ Hybridization.- In Situ Polymerase Chain Reaction.- Results.- Discussion.- 8. Pathogenetic Mechanisms of Animal Orbiviruses That Cause Disease at Low Copy Number.- Materials and Methods.- Experimental Design.- Clinical Samples.- Virus Isolation.- Immunochemistry.- Nested Reverse Transcription Polymerase Chain Reaction.- In Situ Hybridization.- Reverse Transcription In Situ Polymerase Chain Reaction.- Results.- Clinicopathologic Features.- Isolation and Typing of Viruses.- Tissue Distribution and Cellular Localization of Viral Proteins and Nucleic Acids.- Discussion.- 9. Localization of Hepatitis C Virus Using Reverse Transcriptase Fluorescence In Situ 5’ Nuclease Assay.- Determining the Localization Method.- Detection of Gene Expression in Specific Cell Types.- 10. Simultaneous Quantification and Localization of HIV gag-pol Expression Using Ultrasensitive Fluorescence In Situ Hybridization.- 11. Quantification and Localization of Gene Expression: Future Directions.

Dettagli dell'edizione

• Editore: Springer Basel AG
• Data di pubblicazione: 2000
• Lingua: Inglese
• ISBN 10: 0817640347
• ISBN 13: 9780817640347
• Rilegatura: Copertina rigida
• Numero di pagine: 172
• Redattore: Patterson Bruce K.
• Contatto del produttore: non disponibile
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