This volume explores the latest methods used by researchers to study different trypanosomatid parasites. These methods cover numerous disciplines, from organismal biology to molecular mechanism. The chapters in this book cover topics such as high-throughput sequencing; next-generation analysis of trypanosomatid genome stability and instability; DNA repair in cell extracts; ribosome profiling; and the use of CRISPR/Cas9 technology for gene editing. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the…mehr
This volume explores the latest methods used by researchers to study different trypanosomatid parasites. These methods cover numerous disciplines, from organismal biology to molecular mechanism. The chapters in this book cover topics such as high-throughput sequencing; next-generation analysis of trypanosomatid genome stability and instability; DNA repair in cell extracts; ribosome profiling; and the use of CRISPR/Cas9 technology for gene editing. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.
Cutting-edge and practical, Trypanosomatids: Methods and Protocols is a valuable resource for any researcher working with trypanosomatids and trypanosomatid-borne diseases.
Chapters 14, 15, 16, 23, 24, 30, and 48 are open access under a CC BY 4.0 license.
Field Isolation and Cultivation of Trypanosomatids from Insects.- Culturing and Transfection of Pleomorphic Trypanosoma brucei.- In Vitro Culture for Differentiation Simulation of Leishmania spp..- Tsetse Fly Transmission Studies of African Trypanosomes.- Infecting Triatomines with Trypanosomes.- High-Throughput Sequencing for Trypanosome Transcriptome Characterization.- Polysome Profiling and Metabolic Labelling Methods to Measure Translation in Trypanosoma brucei.- Immunoprecipitation for the Analysis of Macromolecular Complexes in Trypanosoma cruzi.- Analysis of the In Vivo Translation Process in Trypanosoma cruzi using Ribosome Profiling.- Proteome-Wide Quantitative Phosphoproteomic Analysis of Trypanosoma brucei Insect and Mammalian Lifecycle Stages.- Genome-Wide Proteomics and Phosphoproteomics Analysis of Trypanosoma cruzi during Differentiation.- Genome-Wide Proteomics and Phosphoproteomics Analysis of Leishmania spp. during Differentiation.- CRISPR/Cas9 Technology Applied to the Study of Proteins Involved in Calcium Signaling in Trypanosoma cruzi.- Application of CRISPR/Cas9 Mediated Genome Editing in Leishmania.- Next Generation Analysis of Trypanosomatid Genome Stability and Instability.- Transcription Factor Analysis in Trypanosomatids.- Identifying Trypanosome Protein-RNA Interactions using RIP-Seq.- The Tethering Assay: A Simple Method for the Characterization of mRNA-Fate Regulators.- RNA-Binding Proteins and their Targets in Trypanosoma brucei: Single Nucleotide Resolution using iCLIP and iCLAP.- In Vivo Tethering System to Isolate RNA-Binding Proteins Regulating mRNA Decay in Leishmania.- Forward Genetics in African Trypanosomes.- Analysis of Base Excision and Single Strand Break Repair Activities in Trypanosomatid Extracts.- Light Microscopy in Trypanosomes: Use of Fluorescent Proteins and Tags.- ImageJ for Partially and Fully Automated Analysis of Trypanosome Micrographs.- Motility Analysis of Trypanosomatids.- Electron Microscopy Techniques Applied to Symbiont-Harboring Trypanosomatids: The Association of the Bacterium with Host Organelles.- AiryScan Super-Resolution Microscopy to Study Trypanosomatid Cell Biology.- All You Ever Wanted to Know About APOL1 and TLFs and Didn't Dare Ask.- Isolation of Leishmania Promastigote Flagella.- Gel Based Methods for the Investigation of Signal Transduction Pathways in Trypanosoma brucei.- Methods to Investigate Signal Transduction Pathways in Trypanosoma cruzi: Cyclic Nucleotide Phosphodiesterases Assay Protocols.- Methods for the Investigation of Trypanosoma cruzi Amastigote Proliferation in Mammalian Host Cells.- Isolation of Extracellular Vesicles from Leishmania spp..- 2D Gel Electrophoresis Analysis of Leishmania Proteomes.- Analysisof the Physiological and Metabolic State of Leishmania using Heavy Water Labelling.- A Scalable Purification Method for Mitochondria from Trypanosoma brucei.- Isolation of Glycosomes from Trypanosoma cruzi.- Sorting the Muck from the Brass: Analysis of Protein Complexes and Cell Lysates.- Measurement of Energy States of the Trypanosomatid Mitochondrion.- Isolation and Characterization of Acidocalcisomes from Trypanosomatids.- Metabolic Control Analysis for Drug Target Prioritization in Trypanosomatids.- Establishment of a Standardized Vaccine Protocol for the Analysis of Protective Immune Responses during Experimental Trypanosome Infections in Mice.- Isolation of Trypanosoma brucei brucei Infection-Derived Splenic Marginal Zone B Cells Based on CD1dHigh/B220High Surface Expression in a Two-Step MACS-FACS Approach.- Cellular Markers for the Identification of Chemo-Resistant Isolates in Leishmania.- In Vitro Image-Based Assay for Trypanosoma cruzi Intracellular Forms.- In Vitro Drug Efficacy Testing against Trypanosoma brucei.- In Vitro Growth Inhibition Assays of Leishmania spp.- In Vivo Bioluminescence Imaging to Assess Compound Efficacy against Trypanosoma brucei.
Field Isolation and Cultivation of Trypanosomatids from Insects.- Culturing and Transfection of Pleomorphic Trypanosoma brucei.- In Vitro Culture for Differentiation Simulation of Leishmania spp..- Tsetse Fly Transmission Studies of African Trypanosomes.- Infecting Triatomines with Trypanosomes.- High-Throughput Sequencing for Trypanosome Transcriptome Characterization.- Polysome Profiling and Metabolic Labelling Methods to Measure Translation in Trypanosoma brucei.- Immunoprecipitation for the Analysis of Macromolecular Complexes in Trypanosoma cruzi.- Analysis of the In Vivo Translation Process in Trypanosoma cruzi using Ribosome Profiling.- Proteome-Wide Quantitative Phosphoproteomic Analysis of Trypanosoma brucei Insect and Mammalian Lifecycle Stages.- Genome-Wide Proteomics and Phosphoproteomics Analysis of Trypanosoma cruzi during Differentiation.- Genome-Wide Proteomics and Phosphoproteomics Analysis of Leishmania spp. during Differentiation.- CRISPR/Cas9 Technology Applied to the Study of Proteins Involved in Calcium Signaling in Trypanosoma cruzi.- Application of CRISPR/Cas9 Mediated Genome Editing in Leishmania.- Next Generation Analysis of Trypanosomatid Genome Stability and Instability.- Transcription Factor Analysis in Trypanosomatids.- Identifying Trypanosome Protein-RNA Interactions using RIP-Seq.- The Tethering Assay: A Simple Method for the Characterization of mRNA-Fate Regulators.- RNA-Binding Proteins and their Targets in Trypanosoma brucei: Single Nucleotide Resolution using iCLIP and iCLAP.- In Vivo Tethering System to Isolate RNA-Binding Proteins Regulating mRNA Decay in Leishmania.- Forward Genetics in African Trypanosomes.- Analysis of Base Excision and Single Strand Break Repair Activities in Trypanosomatid Extracts.- Light Microscopy in Trypanosomes: Use of Fluorescent Proteins and Tags.- ImageJ for Partially and Fully Automated Analysis of Trypanosome Micrographs.- Motility Analysis of Trypanosomatids.- Electron Microscopy Techniques Applied to Symbiont-Harboring Trypanosomatids: The Association of the Bacterium with Host Organelles.- AiryScan Super-Resolution Microscopy to Study Trypanosomatid Cell Biology.- All You Ever Wanted to Know About APOL1 and TLFs and Didn't Dare Ask.- Isolation of Leishmania Promastigote Flagella.- Gel Based Methods for the Investigation of Signal Transduction Pathways in Trypanosoma brucei.- Methods to Investigate Signal Transduction Pathways in Trypanosoma cruzi: Cyclic Nucleotide Phosphodiesterases Assay Protocols.- Methods for the Investigation of Trypanosoma cruzi Amastigote Proliferation in Mammalian Host Cells.- Isolation of Extracellular Vesicles from Leishmania spp..- 2D Gel Electrophoresis Analysis of Leishmania Proteomes.- Analysisof the Physiological and Metabolic State of Leishmania using Heavy Water Labelling.- A Scalable Purification Method for Mitochondria from Trypanosoma brucei.- Isolation of Glycosomes from Trypanosoma cruzi.- Sorting the Muck from the Brass: Analysis of Protein Complexes and Cell Lysates.- Measurement of Energy States of the Trypanosomatid Mitochondrion.- Isolation and Characterization of Acidocalcisomes from Trypanosomatids.- Metabolic Control Analysis for Drug Target Prioritization in Trypanosomatids.- Establishment of a Standardized Vaccine Protocol for the Analysis of Protective Immune Responses during Experimental Trypanosome Infections in Mice.- Isolation of Trypanosoma brucei brucei Infection-Derived Splenic Marginal Zone B Cells Based on CD1dHigh/B220High Surface Expression in a Two-Step MACS-FACS Approach.- Cellular Markers for the Identification of Chemo-Resistant Isolates in Leishmania.- In Vitro Image-Based Assay for Trypanosoma cruzi Intracellular Forms.- In Vitro Drug Efficacy Testing against Trypanosoma brucei.- In Vitro Growth Inhibition Assays of Leishmania spp.- In Vivo Bioluminescence Imaging to Assess Compound Efficacy against Trypanosoma brucei.
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