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In Living Color
Protocols in Flow Cytometry and Cell  Sorting

TABLE OF CONTENTS

INTRODUCTION: A Palette of Living Colors; Rochelle Diamond

HISTORY: Susan DeMaggio

GETTING STARTED

  • Chapter 1: How the FACScalibur and FACSvantage Work and Why It Matters; Tom Frey, Ph.D.
  • Chapter 2:  How Flow Cytometers Work and Don’t Work; Howard M. Shapiro, M.D., P.C.
  • Chapter 3:  Flow Cytometry Standard (FCS) Data File Format; Larry Seamer
  • Chapter 4:  Data Display; Susan DeMaggio and Rochelle Diamond
  • Chapter 5:  Guidelines to Improve Flow Cytometry Data Display and Interpretation;Wendy Schober and Dorothy Lewis
  • Chapter 6:  Data Management and Storage; Dave Coder
  • Chapter 7:  Coloring up: A guide to Spectral Compensation; Rochelle Diamond
  • Chapter 8:  Consistency in Quality Control; Rochelle Diamond


SAMPLE PREPARATION AND CELL SURFACE STAINING

  • Chapter 1:  Cell Preparation and Enrichment for FCM Analysis and Cell Sorting; Rochelle Diamond, Hua Wang, Fei Chen, and Mindy Wilke-Douglas
  • Chapter 2:  The Basics of Staining for Cell Surface Proteins; Alice Givan
  • Chapter 3:  Phenotypic Analysis Using Five-Colour Immunofluorescence and Flow Cytometry; Andrew Beavis
  • Chapter 4:  Detection of Cell Surface Antigens Using Antibody-Conjugated Fluorospheres; Andrew Beavis
  • Chapter 5:  Gel Microdrop Encapsulation for the Frugal Investigator; David McFarland and Gary Durack


REPORTER GENES AND CELL TRACKERS

  • Chapter 1:  Flow Cytometric Analysis of GFP Expression in Mammalian Cells; Steven R. Kain, Ph.D.
  • Chapter 2:  FACS-Gal:  Flow Cytometric Assay of B-galactosidase; Steve Fiering
  • Chapter 3: Bicistronic Vectors and FCM; Nicholas Poulos, Eric Stanbridge, and Susan DeMaggio
  • Chapter 4: NGFR as a Reporter Gene; G. Raghu, M. Munn, S.W. Park, C-H Pan, and A. Dayn
  • Chapter 5:  Assessment of Cell Migration in vivo Using the Fluorescent Tracking Dye PKH26 and Flow Cytometry; Andrew Beavis
  • Chapter 6:  In vivo Biotinylation for Cell Tracking and Survival/Lifespan Measurments; Kenneth A. Ault, M.D. and Cathy Knowles
  • Chapter 7:  MACSelect Chapter – Title still coming; Mindy Wilke-Douglas and Ute Behrens-Jung
  • Chapter 8: Cell Tracking Using PKH; Rebecca Poon and Bruce Bagwell


FLOW CYTOMETRY OF NUCLEIC ACIDS

  • Chapter 1:  RNA Content Determination Using Pyronin Y; Howard Shapiro, M.D., P.C.
  • Chapter 2:  DNA Content Determination; Derek Davies
    • Propidium Iodide Staining for DNA Content
    • Vital Staining for DNA with Hoechst 3342
    • Bromodeoxyuridine Staining for Cell Cycle Kinetics
  • Chapter 3:  Dead Cell Discrimination;  Derek Davies
    • Propidium Iodide Staining for Dead Cell Discrimination
    • Dead Cell Discrimination Using ToPro-3
  • Chapter 4:  Detecting Apoptosis;  Derek Davies & Ingrid Schmid
    • Hoechst/Propidium Iodide Detection for Apoptotic Cells
    • 7-AAD Staining for Apoptosis
    • Annexin-FITC Staining for Apoptotic Cells
  • Chapter 5:  Chromosome Preparation; Derek Davies and Clare Hughes
  • Chapter 6:  D-Flowering – The Flow Cytometry of Plant DNA; John Mottley, K. Yokoya, A.V. Roberts
  • Chapter 7:  Flow Cytometry Analysis of Marine Picoplankton; Dominique Marie, N. Simon, L. Guillon, F. Partensky, D. Vaulot
  • Chapter 8:  Measuring Yeast DNA; Wendy Schober and Jeff Bachant


INTRACELLULAR PHYSIOLOGICAL AND ANTIBODY PROBES

  • Chapter 1:  Calcium Flux in INDO-1 Stained Thymocytes; Geoffrey W. Osborne and Paul Waring, Ph.D.
  • Chapter 2:  Measuring Intracellular pH Using ADB; Mike Fox
  • Chapter 3:  Measuring Intracellular pH Using SNARF-1; Mike Fox
  • Chapter 4:  SNARF-1 and Apoptosis; Judy Norberg, Roberta Gottlieb and Grant Meisenholder
  • Chapter 5:  Measurement of Ligand Acidification Kinetics for Adherant and Non-Adherant Cells; Sheree Lynn Ryback and Robert F. Murphy
  • Chapter 6:  Detection of Intracellular Antigens by Flow Cytometry; Ingrid Schmid
  • Chapter 7:  Flow Cytometric Measurement of  Nuclear Matrix Proteins; Joe Dynlacht
  • Chapter 8:  Flow Cytoenzymology using CellProbe Technology; Susan DeMaggio

ELECTRONIC CELL SORTING

  • Chapter 1:  Creating Standard and Reproducible Sorting Conditions; Marty Bigos, Richard Stoval, and David Parks
  • Chapter 2:  Sterilization for Sorting; Steve Merlin
  • Chapter 3:  High Speed Sorting by MoFlo; Peter Lopez
  • Chapter 4:  Purification of Mouse Fetal Liver Hematopoeitic Stem Cells; Sean Morrison
  • Chapter 5:  Methods for Preparing Sorted Cells for Further Analysis; Robert Lief and Karen Chew

CORE FACILITIES

  • Chapter 1:  Core Facilities for Flow Cytometry; Steve Merlin
  • Chapter 2:  Funding for Core Facilities; Larry Sklar
  • Chapter 3:  Biosafety in the Flow Cytometry Laboratory; Ingrid Schmid


UNLOCKING THE SPECTRUM OF POSSIBILITIES; Rochelle Diamond

APPENDIX

  • Lasers; Paul Ginouves
  • Table of Fluorescent Compounds, Applications, Excitation and Emission spectra, Laser Lines, and Filter Setups; Steve Merlin
  • Common Filter Setup for Large Sorter System – FACS Vantage; Steve Merlin
  • Schematic for following Fluidics Problems;  Steve Merlin, Rebecca Poon
  • PHK  Cell Linker Dyes; Steve Merlin, Rebecca Poon
  • Cell Census Software; Rebecca Poon
  • Monoclonal Antibody Production; Gary Woo
  • Commercial Sources; VARIOUS Authors
  • Networking – Electronic and Personal
    Glossary

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