Methods for Studying Mononuclear Phagocytes

Methods for Studying Mononuclear Phagocytes is a practical guide to the study of mononuclear phagocytes that brings together various well-established and useful methods for examining these cells. The technical protocols have been made detailed, specific, practical, and inclusive of the necessary mystique for immediate and direct application in the laboratory.
The book is divided into 11 parts arranged according to the sequence of steps that would generally be followed to study a given population of mononuclear phagocytes: (I) methods for obtaining and culturing populations of human and animal mononuclear phagocytes; (II) methods for separating populations of leukocytes to enrich or deplete their content of mononuclear phagocytes; (II) criteria and techniques for identifying mononuclear phagocytes; (IV) methods for quantifying the number of mononuclear phagocytes; (V) techniques for studying the morphology of mononuclear phagocytes; (VI) methods for quantifying the biochemical constituents of mononuclear phagocytes; (VII) methods of quantifying phagocytosis, pinocytosis, and chemotaxis; (VIII) methods for quantifying the secretory products of mononuclear phagocytes; (IX) procedures for quantifying the destruction of tumor cells and of microorganisms by mononuclear phagocytes; (X) methods for studying the cell biology of mononuclear phagocytes; and (XI) techniques for studying mononuclear phagocytes in vivo, including procedures for estimating their kinetics, accumulation, identification, and microbicidal properties.

Contenu

Contributors
Preface
I. Obtaining and Culturing Mononuclear Phagocytes
1 The Culture of Mononuclear Phagocytes: A Brief Overview
2 Murine Mononuclear Phagocytes from Bone Marrow
3 Obtaining and Culturing Murine Monocytes
4 Obtaining and Culturing Human Monocytes
5 Isolation of Human Monocytes
6 Culture of Human Monocytes in Microplates and Enzymatic Assays for Following Their Maturation
7 Peritoneal Mononuclear Phagocytes from Small Animals
8 Obtaining and Culturing Human and Animal Alveolar Macrophages
9 Obtaining Human Mononuclear Phagocytes from Colostrum
10 Obtaining Adherent Cells from Spleen
11 Obtaining Kupffer Cells
12 Obtaining Mononuclear Phagocytes from Disaggregated Neoplasms
13 Obtaining Mononuclear Phagocytes from Granulomas
14 Teflon Film as a Substrate for the Culture of Mononuclear Phagocytes
15 Effects of Husbandry and Mouse Strains on Mononuclear Phagocytes
16 Endotoxin Contamination and in Vitro Monocyte-Macrophage Function: Methods of Detecting, Detoxifying, and Eliminating Endotoxin
17 Continuous Cell Lines with Properties of Mononuclear Phagocytes
II. Separation of Mononuclear Phagocytes for Enrichment or Depletion
18 Separation of Mononuclear Phagocytes for Enrichment and for Depletion: An Overview
19 Separation of Murine Macrophages by Adherence to Solid Substrates
20 Separation of Human Monocytes and Guinea Pig Macrophages by Density Gradients of Metrizamide
21 Separation of Murine Mononuclear Phagocytes by Density Gradients of Percoll
22 Separation of Human Monocytes by Density Gradients of Percoll
23 Separation of Murine Mononuclear Phagocytes by Velocity Sedimentation at Unit Gravity
24 Use of Lidocaine for Detachment of Adherent Mononuclear Phagocytes
25 Use of Sephadex Columns to Deplete Mononuclear Phagocytes
26 Depletion of Mononuclear Phagocytes: Pitfalls in the Use of Carbonyl Iron, Carrageenan, Silica, Trypan Blue, or Anti-Mononuclear Phagocytes Serum
III. Identification of Mononuclear Phagocytes
27 Identification of Mononuclear Phagocytes: Overview and Definitions
28 Characteristics of Mononuclear Phagocytes from Different Tissues
29 Fc and C3 Receptors
30 Identification of Mononuclear Phagocytes by Ingestion of Particulate Materials, such as Erythrocytes, Carbon, Zymosan, or Latex
31 Heteroantisera Raised against Mononuclear Phagocytes
32 Monoclonal Antibodies as Tools for the Study of Mononuclear Phagocytes
33 Antisera against Ia Antigens
IV. Quantitation of Number of Mononuclear Phagocytes
34 Quantitation of Adherent Mononuclear Phagocytes by Inverted Phase Microscopy
35 Quantitation of DNA in Mononuclear Phagocytes
36 Lowry and Bradford Assays for Protein
V. Morphology of Mononuclear Phagocytes
37 Use of Wrights Strain and Cytocentrifuge Preparations
38 Use of Phase Contrast Microscopy
39 Use of Peroxidase Stain by the Kaplow Method
40 Use of Nonspecific Esterase Stain
41 Histochemical Stains for Macrophages in Cell Smears and Tissue Sections: ß-Galactosidase, Acid Phosphatase, Nonspecific Esterase, Succinic Dehydrogenase, and Cytochrome Oxidase
42 Use of Transmission Electron Microscopy
43 Preparative Techniques for Scanning Electron Microscopy
44 Use of Ultrastructural Histochemistry
VI. Biochemical Constituents of Mononuclear Phagocytes
45 Lysosomal Enzymes
46 Microsomal Heme Oxygenase
47 Histamine O-Methyltransferase
48 5'-Nucleotidase Assay
49 Alkaline Phosphodiesterase I
50 Quantitation of Leucine Aminopeptidase of Mononuclear Phagocytes
51 Hexose Monophosphate Shunt Activity and Oxygen Uptake
52 Secretion of Superoxide Anion
53 Release of Hydrogen Peroxide
VII. General Functions of Mononuclear Phagocytes
54 Antibody-Dependent and Antibody-Independent Phagocytosis
55 Pinocytic Rate Using Horseradish Peroxidase
56 Chemotaxis of Human and Murine Mononuclear Phagocytes
VIII. Secretion by Mononuclear Phagocytes
57 Secretory Functions of Mononuclear Phagocytes: Overview and Methods for Preparing Conditioned Supernatants
58 Characterization and Classification of Macrophage Proteinases and Proteinase Inhibitors
59 Growth of Macrophage on Collagen, Elastin, and Glycoprotein-Coated Plates as a Tool for Investigating Macrophage Proteinases
60 Neutral Proteases by 3H-Labeled Casein
61 Plasminogen Activators by Use of 3H-Labeled Casein Substrate
62 Elastinolytic Enzymes
63 Microtiter Assay for Antiviral Effects of Human and Murine Interferon Utilizing a Vertical Light Path Photometer for Quantitation
64 Endogenous Pyrogen
65 Prostaglandins
66 Quantitation of Selected Complement Components
67 Lysozyme
IX. Destruction by Mononuclear Phagocytes
68 Destruction of Listeria monocytogenes in Vitro
69 Ingestion and Destruction of Candida albicans
70 Quantitation of Destruction of Toxoplasma
71 Destruction of Rickettsiae
72 Destruction of Leishmania
73 Destruction of Viruses
74 Cytostasis of Tumor and Nontumor Cells
75 Cytolysis of Tumor Cells by Release of [3H] Thymidine
76 Quantification of Cytolysis of Neoplastic Cells by Release of Chromium-51
77 Assessment of Cytolysis of Tumor Cells by Release of [125I] Iododeoxyuridine
78 Antibody-Dependent Cellular Cytotoxicity (ADCC) of Erythroid and Tumor Cells
X. Mononuclear Phagocytes as Tools in Cell Biology
79 Overview: The Macrophage in Cell Biology
80 Biosynthetic Radiolabeling of Cellular and Secreted Proteins of Mononuclear Phagocytes
81 Extraction, Identification, and Quantitation of Lipids
82 Synthesis, Cellular Turnover, and Mass of Cholesterol
83 Solute Uptake and Membrane Transport by Mononuclear Phagocytes
84 Use of Lactoperoxidase for Labeling Membrane Proteins
85 Binding of Synthetic Chemotactic Peptides as a Model of Ligand-Receptor Interaction
XI. Mononuclear Phagocytes In Vivo
86 Isolation of Phagosomes from Mouse Peritoneal Macrophages
87 Study of Mononuclear Macrophages in Vivo: Overview
88 Quantitation of the Inflammatory Accumulation of Mononuclear Phagocytes in Vivo
89 Systemic Labeling of Mononuclear Phagocytes
90 Labeling of Mononuclear Phagocytes in Granulomas and Inflammatory Sites
91 Identification of Fc and Complement Rec…