Cell Counting

Mary C. Phelan1, Gretchen Lawler2

1 Thompson Children's Hospital, Chattanooga, Tennessee, 2 Purdue University, West Lafayette, Indiana
Publication Name:  Current Protocols in Cytometry
Unit Number:  Appendix 3A
DOI:  10.1002/0471142956.cya03as00
Online Posting Date:  May, 2001
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Abstract

This unit presents protocols for counting cells using either a hemacytometer or electronically using a Coulter counter. Cell counting with a hemacytometer permits effective discrimination of live from dead cells using trypan blue exclusion. In addition, the procedure is less subject to errors arising from cell clumping or size heterogeneity. Counting cells is more quickly and easily performed using an electronic counter, but live‚Äźdead discrimination is unreliable. Cell populations containing large numbers of dead cells and/or cell clumps are difficult to count accurately. In addition, electronic counting requires resetting of the instrument for cell populations of different sizes; heterogeneous populations can give rise to inaccurate counts, and resting and activated cells may require counting at separate settings. In general, electronic cell counting is best performed on fresh peripheral blood cells.

     
 
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Table of Contents

  • Basic Protocol 1: Counting Cells Using a Hemacytometer
  • Basic Protocol 2: Counting Cells Using a Coulter Counter
  • Commentary
  • Figures
     
 
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Materials

Basic Protocol 1: Counting Cells Using a Hemacytometer

  Materials
  • 70% (v/v) ethanol
  • Cell suspension
  • 0.4% (w/v) trypan blue or 0.4% (w/v) nigrosin, prepared in HBSS ( appendix 2A)
  • Hemacytometer with coverslip (e.g., Improved Neubauer, VWR)
  • Hand‐held counter

Basic Protocol 2: Counting Cells Using a Coulter Counter

  Materials
  • Cell suspension
  • 20 ml counting vials
  • Zap‐O‐globin (Coulter), lysing and hemoglobin reagent (VWR), or equivalent (for counting white blood cells)
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Figures

Literature Cited

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