Full Description
Neutrophils, the most abundant white cells in humans, serve as the primary cellular defense against infection. Neutrophil Methods and Protocols, Second Edition provides a concise set of protocols written by leading researchers in the field for assessing basic neutrophil functions, investigating specialized areas in neutrophil research, and completing step diagnostic assays of common neutrophil disorders. Topics covered include an overview of neutrophils and their role in host defense and inflammation; methods most commonly used for isolating neutrophils from humans and other animal species; procedures for subcellular fractionation of human neutrophils, analysis of in vivo transmigrated neutrophils, generation of mature neutrophils from induced pluripotent stem cells and analysis of neutrophil gene expression; methods for investigating priming, oxidant production, phagocytosis, bactericidal activity and extracellular trap formation and protocols for investigating neutrophil adhesion, chemotaxis and outside-in signaling via integrins. Written in the 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 protocols, and notes on troubleshooting and avoiding known pitfalls.
Authoritative and easily accessible, Neutrophil Methods and Protocols, Second Edition is a comprehensive source for detailed explanations and applications of the most modern methodological advances in neutrophil biology. Both basic scientists and clinicians will find a collection of this caliber to be an invaluable aid in their work with neutrophils.
Contents
The Role of Neutrophils in the Immune System: An Overview.- Isolation of Human Neutrophils.- Neutrophil Isolation from Non-Human Species.- Collection of In vivo Transmigrated Neutrophils from Human Skin.- Subcellular Fractionation of Human Neutrophils and Analysis of Subcellular Markers.- Rho Family and Rap GTPase Activation Assays.- Measurement of Phospholipid Metabolism in Intact Neutrophils.- Optical Methods for the Measurement and Manipulation of Cytosolic Free Calcium Signals in Neutrophils.- Analysis of Electrophysiological Properties and Responses of Neutrophils.- Analysis of Neutrophil Apoptosis.- Microinjection Methods for Neutrophils.- Generation of Functionally Mature Neutrophils from Induced Pluripotent Stem Cells.- Neutrophil Migration Through Extracellular Matrix.- Spinning Disk Confocal Imaging of Neutrophil Migration in Zebrafish.- Detection of Bi-Directional Signalilng During Integrin Activation and In Neutrophil Adhesion.- Immunofluorescence and Confocal Microscopy of Neutrophils.- Expression of Genetically-Encoded Fluorescent Probes to Monitor Phospholipid Dynamics in Live Neutrophils.- Quantitative Assessment of Neutrophil Phagocytosis Using Flow Cytometry.- Analysis of Neutrophil Bactericidal Activity.- Induction and Quantification of Neutrophil Extracellular Traps.- Measurement of Respiratory Burst Products, Released or Retained, During Activation of Professional Phagocytes.- Cell-free NADPH Oxidase Assays: "In vitro Veritas".- Assessment of Priming of the Human Neutrophil Respiratory Burst.- Affinity Purification and Reconstitution of Human Phagocyte Flavocytochrome b for Detection of Conformational Dynamics in the Membrane.- Evaluation of p47phox Phosphorylation in Human Neutrophils Using Phospho-Specific Antibodies.- Genome-Scale Transcript Analyses with Human Neutrophils.- Fast and Accurate Quantitative Analysis of Cytokine Gene Expression in Human Neutrophils.- High Purity Neutrophil Isolation from Human Peripheral Blood and Saliva for Transcriptome Analysis.- Detection of Intact Transcription Factors in Human Neutrophils.- Disorders of Neutrophil Function: An Overview.- Diagnostic Assays for Chronic Granulomatous Disease and Other Neutrophil Disorders.- Diagnostic Assays for Myeloperoxidase and Myeloperoxidase Deficiency.
