Platelets (thrombocytes) are small, irregularly shaped clear cell fragments derived from megakaryocytes and are necessary for normal blood clotting. Platelets, by nature, are very sticky, and when activated, will adhere easily to other sticky cell types (i.e. monocytes, eosinophils, etc.), creating cell clumps.
Follow these tips to reduce platelet contamination and prevent platelet activation in your blood samples.
Recommendations for Blood Sample Collection
Samples containing platelets should be collected and handled gently to prevent platelet activation. The following recommendations1 may help:
When drawing blood, use a wide bore needle (e.g. 19G), and do not apply vigorous traction if using a syringe plunger.
Use anticoagulants when drawing blood (e.g. collect the blood in acid citrate dextrose (ACD) tubes).
Do not vortex the blood sample.
Use buffer without Ca++ and Mg++ to prevent platelet activation.
Preparation of Mononuclear Cell Suspensions
The following recommendations may help reduce platelet contamination in mononuclear cell suspensions prepared from fresh whole blood:
When isolating mononuclear cells during density gradient centrifugation (e.g. using Lymphoprep™ or Ficoll™ as the density medium), first pipette off the upper, platelet-rich plasma layer as much as possible without disturbing the PBMC layer.
To wash the isolated mononuclear cells:
Perform a slow spin on the isolated cells (120 x g, 10 min, brake off, room temperature).
Ensure the centrifuge swinging-buckets and the tubes inside are perfectly balanced. Since this is a low g-force spin, even small imbalances may dislodge the pellet.
Carefully remove the platelet-rich supernatant and discard.
Using a fresh pipette, gently resuspend the cell pellet in D-PBS with 2% FBS.
Repeat at least twice for a total of 3 or more washes.
The standard procedure for the isolation of peripheral blood mononuclear cells (PBMCs) using density gradient centrifugation does not remove platelets. Researchers can perform additional washing steps in order to remove platelets following density gradient centrifugation. Alternatively, researchers can use the EasySep™ Direct Human PBMC Isolation Kit, which immunomagnetically depletes platelets during PBMC isolation. See the data >
Isolating Target Cells Using Magnetic Cell Separation
STEMCELL Technologies also provides a platelet removal cocktail (Catalog# 19369C) to further deplete platelets during cell isolation using the EasySep™ Isolation or Enrichment kits. For more information on this product or to discuss compatibility with your cell isolation system, please contact firstname.lastname@example.org.
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