Flow Cytometry: A Practical ApproachM. G. Ormerod OUP Oxford, 18 de mai. de 2000 - 276 páginas Flow Cytometry 3/e is intended as a handbook for every laboratory that has a bench-top flow cytometer or a fluorescence activated cell sorter. It is an introduction and guide to those new to the field and a first point of reference for experienced practitioners who want to investigate a new technique. The chapter on immunophenotyping - the most important clinical application of flow cytometry - has been strengthened by the addition of a chapter on quality control in the clinical laboratory. The utility of the book in a clinical laboratory has been further enhanced by the addition of a chapter covering ten other clinical applications. Flow cytometry has found increasing application in the field of apoptosis research. A new chapter has been added to cover this important topic. Every flow cytometry laboratory can't afford not to have a copy on the shelf as a first point of reference. The book is not fully comprehensive but it does aim to cover over 90% of the applications of flow cytometry in mammalian biology. |
Conteúdo
Introduction to the principles of flow cytometry | 1 |
2 Techniques for sample preparation | 2 |
3 Fluidics | 3 |
4 Detection and measurement | 7 |
5 Cell sorting | 21 |
References | 22 |
Fluorescence and fluorochromes | 23 |
3 Fluorochromes used to label covalently other probes | 25 |
References | 132 |
Measurement of cytoplasmic and nuclear antigens | 133 |
2 Methods of cell permeabilization | 134 |
3 Methods of cell staining and analysis | 142 |
4 Examples of intracellular antigens that can be measured by flow cytometry | 146 |
References | 155 |
Analysis of DNAmeasurement of cell kinetics by the bromodeoxyuridineantibromodeoxyuridine method | 159 |
2 Basic cellkinetic concepts | 160 |
4 Fluorochromes used to label directly cell components | 28 |
References | 33 |
Preparing suspensions of single cells | 35 |
3 Leucocytes from other tissue | 38 |
4 Other types of cell | 39 |
5 Cultured cells | 44 |
References | 45 |
Flow sorting | 47 |
3 Parameters which affect sorting | 50 |
4 Practical considerations | 52 |
5 Specialized features | 57 |
6 Other flowsorting systems | 58 |
Further reading | 59 |
Immunofluorescence of surface markers | 61 |
2 Instrument standardization | 63 |
3 Testing sensitivity and resolution | 68 |
4 Immunofluorescence | 70 |
6 Titration of antibodies | 72 |
9 Constructing an immunophenotyping panel | 73 |
10 Isotype controls | 80 |
11 DNA contentsurface marker analysis | 81 |
References | 82 |
Analysis of DNAgeneral methods | 83 |
3 General comments | 85 |
4 Experimental methods | 89 |
5 Deconvolution of the DNA histogram | 93 |
Acknowledgement | 96 |
Further clinical applications | 99 |
3 The enumeration of CD34+ cells in peripheral blood stemcell harvests and bonemarrow harvests | 101 |
4 Simultaneous detection of granulocyte and lymphocytereactive antibodies | 104 |
5 Detection of plateletreactive antibodies | 107 |
6 Application of crossmatching by flow cytometry to transplantation | 109 |
7 Monitoring antithymocyte globulin ATGantilymphocyte globulin ALG therapy in transplantation | 111 |
8 Measurement of cellmediated cytotoxicity by flow cytometry | 114 |
9 Monitoring cell activation in response to antigens or mitogens | 116 |
10 Measuring intracellular cytokines by flow cytometry | 117 |
References | 122 |
Quality assurance in the clinical laboratory | 125 |
3 Internal qualitycontrol procedures | 127 |
4 External quality assurance | 130 |
5 Conclusions | 131 |
3 Background to the BrdUrd technique | 162 |
4 Methods of BrdUrd incorporation | 164 |
5 Tissue preparation | 166 |
6 Staining procedures | 167 |
7 Flow cytometry | 172 |
9 Conclusions | 176 |
Acknowledgements | 177 |
Analysis of cell proliferation using the bromodeoxyuridineHoechstethidium bromide method | 179 |
3 Cell staining and flow cytometry | 180 |
4 Synchronized cells | 182 |
5 Asynchronous cells | 183 |
6 Troubleshooting | 186 |
Acknowledgements | 188 |
Chromosome analysis and sorting by flow cytometry | 189 |
2 Preparation of chromosome suspensions | 192 |
3 Instrumentation | 196 |
4 Flow sorting chromosomes for library construction | 197 |
5 Generation of chromosome paints | 198 |
References | 201 |
Intracellular ionized calcium magnesium membrane potential and pH | 203 |
2 Magnesium | 222 |
4 Measurement of cytoplasmic pH | 227 |
Acknowledgement | 232 |
Flow cytometry in the study of apoptosis | 235 |
3 Measurement of DNA degradation | 236 |
4 Mitochondrial membrane potential | 240 |
5 Measurements of changes in the plasma membrane | 243 |
6 Other measurements | 245 |
7 Quantification of apoptotic cells | 246 |
Acknowledgement | 247 |
Further applications to cell biology | 249 |
3 Monitoring electropermeabilization of cells | 251 |
4 Measurement of oxidative burst | 253 |
5 Characterizing multidrug resistance MDR in cancer cells | 254 |
Safety procedures | 259 |
3 Chemicals | 260 |
Flow cytometers and software | 261 |
2 Software for data manipulation | 263 |
List of suppliers | 265 |
273 | |
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Termos e frases comuns
analysis antigens apoptosis apoptotic apoptotic cells assay Becton Dickinson binding BrdUrd buffer Ca2+ calcium cell cycle cell line cell suspension cellular Centrifuge channel Chapter chromosomes clinical concentration conjugate culture cytogram cytokines cytometer cytoplasmic detection diluted display DNA content DNA histogram droplet emission Equipment and reagents ethanol excitation Figure filter FITC fixation flow chamber flow cytometry fluorescein fluorescence fluorochromes Fura Red G₁ G₂ Hoechst immunofluorescence Immunol immunophenotyping Incubate instrument intracellular antigens isotype control labelled laboratory leucocytes light scatter light-scatter gate lymphocytes markers measured membrane potential method Molecular Probes monoclonal antibodies monocytes nuclei number of cells optical Ormerod parameters particles PCNA peak peripheral blood permeabilization phase platelets plot preparation procedure propidium iodide protein Protocol ratio right-angle room temperature sample Sigma signal single cells SNARF-1 solution sorting staining standard supernatant technique tissue tube tumour voltage Wash wavelength µg/ml