Techniques in Quantification and Localization of Gene Expression - Tapa blanda

Sinopsis

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.

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