Name: preparation of pooled human platelet lysate (phpl) as an efficient supplement for animal serum-free human stem cell cultures
Uploaded: 2009-10-30T07:00:00
Description: Watch this Scientific Journal Video about Preparation of Pooled Human Platelet Lysate pHPL as an Efficient Supplement for Animal Serum-Free Human Stem Cell Cultures at JoVE.com

Ce travail a été soutenu par l&#39;Austrian Research Foundation (FWF, accorder N211-NAN à la DS) et l&#39;Agence autrichienne de promotion de la recherche (FFG, accorder à la N200 DS). Les auteurs remercient Url Claudia pour l&#39;assistance technique excellente et Monica Farrell pour l&#39;édition linguistique.

1. Matériau de départ Commencez par plasma riche en plaquettes (PRP) unités préparées par cytaphérèse ou dérivée de Buffy contre les manteaux. 2. Vérifier la stérilité Pour vérifier la stérilité de prendre un échantillon de 20 ml de chaque unité de PRP en transférant le volume d&#39;un sac de petite connecté (Baxter). Débranchez ce sac par soudage. 3. Gel des unités PRP <p class="jove_co...

<ol>
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J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficacy+and+safety+of+the+use+of+autologous+plasma+rich+in+platelets+for+tissue+regeneration:+a+systematic+review.">Efficacy and safety of the use of autologous plasma rich in platelets for tissue regeneration: a systematic review.</a> Transfusion. 49, 44-56 (2009).</li><li>Weibrich, G., Kleis, W. K., Hafner, G., Hitzler, W. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+factor+levels+in+platelet-rich+plasma+and+correlations+with+donor+age,+sex,+and+platelet+count.">Growth factor levels in platelet-rich plasma and correlations with donor age, sex, and platelet count.</a> J Craniomaxillofac Surg. 30, 97-102 (2002).</li><li>Frechette, J. P., Martineau, I. &. a. m. p. ;. a. m. p., Gagnon, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Platelet-rich+plasmas:+growth+factor+content+and+roles+in+wound+healing.">Platelet-rich plasmas: growth factor content and roles in wound healing.</a> J Dent Res. 84, 434-439 (2005).</li><li>Seghatchian, J., Krailadsiri, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+platelet+storage+lesion.">The platelet storage lesion.</a> Transfus Med Rev. 11, 130-144 (1997).</li><li>Selvaggi, T. A., Walker, R. E., Fleisher, T. 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Dans certaines régions plasma riche en plaquettes (PRP) peuvent être obtenus à partir de buffy-coats contraire être un produit des déchets d&#39;emballage production de globules rouges provenant de dons de sang testés (figure 1). De façon optimale, le PRP est utilisé immédiatement pour la préparation ultérieure d&#39;pHPL, comme en plaquettes périmées concentre la disponibilité de facteurs de croissance peuvent être réduites en raison de lésions de stockage de plaquettes et de la dégradation de 20...

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		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Double bag (2 x 3.5L)</td>
<td>&nbsp;</td>
<td>MacoPharma</td>
<td>VDL 8000XQ</td>
<td>originally for ascites puncture</td>
</tr>
<tr>
<td>50 mL Falcon tube</td>
<td>&nbsp;</td>
<td>Falcon BD</td>
<td>2098</td>
<td>&nbsp;</td>
<tr>
<td>50 mL Stripettes</td>
<td>&nbsp;</td>
<td>Costar</td>
<td>4501</td>
<td>&nbsp;</td>
<tr>
<td>Plasma bags (600mL)</td>
<td>&nbsp;</td>
<td>Baxter</td>
<td>R4R2021</td>
<td>&nbsp;</td>
<tr>
<td>Sterile tubing welder</td>
<td>&nbsp;</td>
<td>Terumo</td>
<td>TSCD-II</td>
<td>&nbsp;</td>
<tr>
<td>Welding equipment </td>
<td>&nbsp;</td>
<td>Fresenius Kabi</td>
<td>Compo Seal</td>
<td>&nbsp;</td>
<tr>
<td>Water bath</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Sterile scissors</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Clamps</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Centrifuge</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

preparation of pooled human platelet lysate (phpl) as an efficient supplement for animal serum-free human stem cell cultures

Facteurs de croissance dérivés des plaquettes ont été montré pour stimuler efficacement la prolifération des cellules<em&gt; In vivo</em<sup&gt; 1,2</sup&gt; Et<em&gt; In vitro</em&gt;. Cet effet a été rapporté pour les cellules stromales mésenchymateuses (CSM), des fibroblastes et endothéliales cellules formant des colonies avec des plaquettes activées par la thrombine<sup&gt; 3-5</sup&gt; Ou lysées par cycles gel / dégel<sup&gt; 6-14</sup&gt; Avant la releasate plaquettaire est ajouté au milieu de culture cellulaire. L&#39;effet trophique des facteurs de croissance plaquettaire dérivés a déjà été testée dans plusieurs essais pour l&#39;ingénierie tissulaire et la thérapie régénérative.<sup&gt; 1,15-17</supEfficacité&gt; Varier est considéré comme au moins en partie due à des concentrations individuellement divergentes des facteurs de croissance<sup&gt; 18,19</sup&gt; Et un manque actuel de protocoles normalisés pour la préparation de plaquettes.<sup&gt; 15,16</sup&gt; Ce protocole présente une procédure possible pour générer un pool de lysat plaquettaire humain (pHPL) dérivé de routine produit un plasma riche en plaquettes (PRP) du 40 au 50 dons de sang unique. Par plusieurs cycles gel / dégel des membranes plaquettaires sont endommagés et les facteurs de croissance sont efficacement libérée dans le plasma. Enfin, les fragments de plaquettes sont éliminées par centrifugation afin d&#39;éviter la formation d&#39;agrégats vaste et épuisent les antigènes potentiels. La mise en œuvre des protocoles de culture en pHPL norme représente un outil prometteur pour le développement de thérapies cellulaires propagé dans un animal sans protéine du système.

Watch this Scientific Journal Video about Preparation of Pooled Human Platelet Lysate pHPL as an Efficient Supplement for Animal Serum-Free Human Stem Cell Cultures at JoVE.com

Preparation of Pooled Human Platelet Lysate pHPL as an Efficient Supplement for Animal Serum-Free Human Stem Cell Cultures

Homme lysat plaquettaire est une riche source de facteurs de croissance et d&#39;un supplément puissant dans la culture cellulaire. Ce protocole présente le processus de préparation d&#39;un grand bassin de lysat plaquettaire humaine en commençant à partir du plasma riche en plaquettes, effectuant plusieurs cycles gel-dégel et d&#39;épuiser les fragments de plaquettes.

Préparation du Pooled lysat plaquettaire humain (pHPL) comme un supplément efficace pour animaux sans sérum humain cultures de cellules souches

Platelet derived growth factors have been shown to stimulate cell proliferation efficiently in vivo1,2 and in vitro. This effect has been reported for ...

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