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This fully updated volume assembles a comprehensive collection of methods, techniques, and strategies to investigate the molecular and cellular biology of peroxisomes in different organisms. Peroxisome research is on the rise, as novel functions and proteins of this dynamic organelle are still being discovered through studies in model systems including humans, mice, flies, plants, fungi, and yeast, and this progress is reflected in the chapters included in this collection. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and up-to-date, Peroxisomes: Methods and Protocols, Second Edition serves as an ideal guide for researchers working on peroxisome- and organelle-based research questions.
Isolation of Mammalian Peroxisomes by Density Gradient Centrifugation
Analysis of Yeast Peroxisomes via Spatial Proteomics
Isolation of Glycosomes from Trypanosoma brucei
Immunolabeling for Detection of Endogenous and Overexpressed Peroxisomal Proteins in Mammalian Cells
Super-Resolution Imaging of Peroxisomal Proteins Using STED Nanoscopy
Direct Stochastic Optical Reconstruction Microscopy (dSTORM) of Peroxisomes
Correlative Light- and Electron Microscopy in Peroxisome Research
Ultrastructural​ Analysis and Quantification of Peroxisome-Organelle Contacts
Detection of Peroxisomal Proteins During Mycobacterial Infection
Proximity-Ligation Assay to Detect Peroxisome-Organelle Interaction
Assay of Reactive Oxygen/Nitrogen Species (ROS/RNS) in Arabidopsis Peroxisomes Through Fluorescent Protein Containing a Type 1 Peroxisomal Targeting Signal (PTS1)
Identification​ of Peroxisome-Derived Hydrogen Peroxide-Sensitive Target Proteins Using a YAP1C-Based Genetic Probe
Assessment of the Peroxisomal Redox State in Living Cells Using NADPH- and NAD /NADH-Specific Fluorescent Protein Sensors
Live-Cell Imaging of Peroxisomal Calcium Levels and Dynamics
Analysis of Peroxisome Biogenesis by Phos-Tag SDS-PAGE
Targeted Modifications of the Yeast Genome to Study Peroxisomes
Applying CRISPR-Cas9 Genome Editing to Study Genes Involved in Peroxisome Biogenesis or Peroxisomal Functions
Generation of Reporter Cell Lines for Endogenous Expression Analysis of Peroxisomal Proteins
Peroxisomes and Viruses:​ Overview on Current Knowledge and Experimental Approaches
Tools to Investigate the Peroxisome-Dependent Antiviral Response
Determining the Importance of Peroxisomal Proteins for Viral Infections in Cultured Mammalian Cells
Characterizati​on of Yeast Peroxisomes:​ Enrichment of Peroxisomal Fractions and Analysis of β-Oxidation Activity
A Cell-Free In Vitro Import System for Peroxisomal Proteins Containing a Type 2 Targeting Signal (PTS2)
Assessing Peroxisomal Protein Interaction by Immunoprecipitat​ion
Purification of a Recombinant Human PEX1/​PEX6 AAA  ATPase Complex from HEK293TT Cells
Affinity Purification of Soluble and Membrane-Bound Protein Complexes by a FlpIn Strategy
Utilization of Nonstop mRNA to Assess Ribosome-Associated Nascent Polypeptide Chains in Early Topogenesis of Peroxisomal Proteins
Computational Evaluation of Peroxisomal Targeting Signals in Metazoa
Computational Approaches for Peroxisomal Protein Localization
Estimating the Interaction Strength Between PTS1-Peptides and Their Receptor PEX5 in Living Cells Using Flow-Cytometer-Based FRET (flowFRET) Measurements
Using the Superfolder GFP (sfGFP) System to Study Plant Peroxisomal Protein Import
Biochemical Fractionation of Trypanosomes for the Analysis of Glycosomal Protein Import Defects
Manipulation and Visualization of Peroxisomes in Drosophila
Mouse Models to Study Peroxisomal Functions and Disorders:​ Overview, Caveats, and Recommendations