Name: counting human neural stem cells
Uploaded: 2007-08-22T00:00:00
Description: Watch this Scientific Journal Video about Counting Human Neural Stem Cells at JoVE.com

<p class="jove_content">The authors would like to acknowledge Dr. Philip H. Schwartz of the National Human Neural Stem Cell Resource at the Children s Hospital, Orange County Research Institute for providing hNSPC cultures.</p>

<p class="jove_step">Passaging Human Neural Stem Cells</p>
<p class="jove_content"><em>Note</em>: Please refer to the <a href="https://www.jove.com/index/details.stp?ID=263"><strong class="jove_content">Passaging Human Neural Stem Cells</strong></a><span class="jove_content"> article on how to detach and resuspend the cells. https://www.jove.com/index/Details.stp?ID=263</span></p>
<p class="jove_step">Preparing the Cell Suspension for Counting</p>
<ol>
<li>Place 25 &micro;l of 0.4% Trypan Blue solution and 20 &micro;l of cell media into an eppe...

<ol>
	<li>Marchenko, S., Flanagan, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Passaging+Human+Neural+Stem+Cells.">Passaging Human Neural Stem Cells.</a> <em style='font-style: italic'>Journal of Visualized Experiments</em>. <b>7</b>, (2007).</li><li>Caprette, D. a. v. i. d. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Using+a+Hemacytometer.">Using a Hemacytometer.</a> <em style='font-style: italic'>BioEd Online</em>. , (2008).</li><li>Caprette, D. a. v. i. d. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Counting+Chamber+(Hemacytometer).">Counting Chamber (Hemacytometer).</a> <em style='font-style: italic'>BioEd Online</em>. , (2008).</li></ol>

<p class="jove_content">This protocol describes a straightforward and rapid way to determine the cell concentration for a variety of experiments involving cells.  Using the shortcut described in 3d, one can quickly and accurately determine the cell concentration and total cell number in a given volume.</p>

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		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
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<tr>
<td>Phase Contrast Hemacytometer</td>
<td>Tool</td>
<td>Hausser </td>
<td>02-671-54</td>
<td>Distributed by Fisher under the indicated catalog #</td>
</tr>
<tr>
<td>Trypan Blue Stain 0.4%</td>
<td>Reagent</td>
<td>Gibco</td>
<td>15250-061</td>
<td>Distributed by Invitrogen under the indicated catalog #</td>
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counting human neural stem cells

<p class="jove_content">Knowledge of the exact number of viable cells in a given volume of a cell suspension is required for many routine tissue culture manipulations, such as plating cells for immunocytochemistry or for cell transfections.  This protocol describes a straightforward and fast method for differentiating between live and dead cells and quantifying the cell concentration and total cell number using a hemacytometer.  This procedure first requires detaching cells from a growth surface and resuspending them in media.  Next, the cells are diluted in a solution of Trypan blue (ideally to a concentration that will give 20-50 cells per quadrant) and placed in the hemacytometer.  Finally, averaging the counts of viable cells in several randomly selected quadrants, dividing the average by the volume of one 1 mm<sup>2</sup> quadrant (0.1  &mu;l) and multiplying by the dilution factor gives the number of cells per  l.  Multiplying this cell concentration by the total volume in  &mu;l gives the total cell number. This protocol describes counting human neural stem/precursor cells (hNSPCs), but can also be used for many other cell types.</p>

Watch this Scientific Journal Video about Counting Human Neural Stem Cells at JoVE.com

Counting Human Neural Stem Cells

<p class="jove_content">Knowledge of the exact number of viable cells is required for many tissue culture manipulations.  This protocol describes how to differentiate between live and dead cells and quantify cells using a hemacytometer.  Although it describes counting human neural stem/precursor cells (hNSPCs), it can be used for other cell types.</p>

Knowledge of the exact number of viable cells in a given volume of a cell suspension is required for many routine tissue culture manipulations, such as ...

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