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

Diese Arbeit wurde von der Austrian Research Foundation (FWF, gewähren N211-NAN DS) und die Österreichische Forschungsförderungsgesellschaft mbH (FFG, gewähren N200 zum DS) unterstützt worden. Die Autoren danken Claudia URL für hervorragende technische Unterstützung und Monica Farrell für die sprachliche Bearbeitung.

1. Ausgangsmaterial Beginnen Sie mit Thrombozyten-reichem Plasma (PRP) Einheiten durch Zytopherese oder aus Buffy Coats. 2. Sterilität zu überprüfen Für die Sterilität zu überprüfen eine Probe von 20 ml aus jedem PRP-Einheit durch die Übertragung der Lautstärke auf einem angeschlossenen kleinen Beutel (Baxter). Trennen Sie diese Tasche durch Schweißen. 3. Einfrieren von PRP-Einheiten <p class="jove...

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In einigen Regionen platelet rich plasma (PRP) kann von Buffy-Coats sonst, ein Abfallprodukt der gepackten roten Blutkörperchen aus getestet Blutspenden (Abbildung 1) erreicht werden. Optimal ist PRP sofort für die weitere Bearbeitung von PHPL verwendet, wie in veralteten Thrombozytenkonzentraten die Verfügbarkeit von Wachstumsfaktoren können durch Thrombozyten-Lagerung Läsionen und Abbau 20 reduziert werden. Es wird ferner empfohlen, PRP durch Anpassung Thrombozyten der Blutgruppe O mit Plasma der Blutg...

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		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
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<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>
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preparation of pooled human platelet lysate (phpl) as an efficient supplement for animal serum-free human stem cell cultures

Platelet derived growth Faktoren haben gezeigt, dass die Zellproliferation effizient stimulieren<em&gt; In vivo</em<sup&gt; 1,2</sup&gt; Und<em&gt; In-vitro-</em&gt;. Dieser Effekt wurde für mesenchymale Stromazellen (MSCs), Fibroblasten und Endothelzellen Kolonie-bildenden Zellen mit Thrombozyten durch Thrombin aktiviert gemeldet<sup&gt; 3-5</sup&gt; Oder durch Frost / Tau-Zyklen lysiert<sup&gt; 6-14</sup&gt; Vor der Thrombozyten releasate ist das Zellkulturmedium aufgenommen. Die trophische Wirkung von thrombozytären Wachstumsfaktoren hat bereits in mehreren Studien für das Tissue Engineering und der regenerativen Therapie getestet.<sup&gt; 1,15-17</sup&gt; Unterschiedliche Effizienz gilt als zumindest teilweise durch individuell unterschiedliche Konzentrationen von Wachstumsfaktoren werden<sup&gt; 18,19</sup&gt; Und einem aktuellen Mangel an standardisierten Protokollen für Thrombozyten-Vorbereitung.<sup&gt; 15,16</sup&gt; Dieses Protokoll stellt ein praktikables Verfahren zu einem Pool von menschlichen Thrombozyten-Lysat (PHPL) von routinemäßig produziert platelet rich plasma (PRP) von 40 bis 50 einzelnen Blutspenden gewonnen zu generieren. Durch mehrere Frost / Tau-Zyklen der Thrombozyten-Membranen sind beschädigt und Wachstumsfaktoren sind effizient in das Plasma freigesetzt. Schließlich sind die Thrombozyten-Fragmenten durch Zentrifugation umfangreiche Aggregatbildung zu vermeiden und zum Abbau potenzieller Antigene entfernt. Die Umsetzung der PHPL in Standard-Kultur-Protokolle stellt ein viel versprechendes Instrument für die weitere Entwicklung von Zelltherapeutika propagiert in einem Tier-Protein-freies System.

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

Menschliche Thrombozyten-Lysat ist eine reiche Quelle von Wachstumsfaktoren und eine starke Ergänzung in der Zellkultur. Dieses Protokoll stellt den Prozess der Vorbereitung einen großen Pool an menschlichen Thrombozyten-Lysat ausgehend von platelet rich plasma, Durchführung mehrerer Gefrier-Auftau-Zyklen und zum Abbau der Thrombozyten-Fragmente.

Vorbereitung der gepoolten menschlichen Thrombozyten-Lysat (PHPL) als effiziente Ergänzung zum tierischen Serum-freien menschlichen Stammzell-Kulturen

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

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