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Laboratory Protocol for Platelet and Serotonin Isolation Using Centrifugation Techniques

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This protocol outlines the step-by-step procedure for isolating platelets and serotonin from blood samples using a centrifugation method. Begin by collecting blood in a 4 mL EDTA tube and centrifuging it for 10 minutes at 200 RCF (xg) at ambient temperature. Remove 400 µL of plasma and transfer it to a 15 mL tube for platelet purification. Following a second centrifugation at 4,500 RCF (xg) and 4°C, vortex the pellet and aliquot the platelets for storage at -80°C. Important factors affecting specimen integrity, such as storage conditions and proper handling, are also discussed.

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Laboratory Protocol for Platelet and Serotonin Isolation Using Centrifugation Techniques

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  1. Platelet Serotonin Laboratory Protocol Centrifuge 10 minutes at 200 RCF(xg) at ambient temperature 4 mL EDTA tube • Remove 400 uL of plasma • Add to 15 mL tube for platelet purification • Add 1.5 mL physiological saline to platelet-rich plasma (PRP) Centrifuge10 minutes at 4,500 RCF(xg) at 4oC & discard supernatant • Add to 300 uL distilled water • Vortex pellet until the platelets are mixed & transfer 300 uL to 1 aliquot ELISA Transfer to -80oC freezer for storage Immuno Biological Laboratories (2009). Serotonin ELISA IB89546. [Laboratory instructions]. IBL-America

  2. Factors Affecting Biological Specimens Specimen Storage Processing Phlebotomy Specimen Transport • Storage temperature • Duration of storage • Freeze-thaw cycles • Instrument calibration & controls • Time between collection & centrifugation • Vortex technique • Centrifugation • Type • Calibrated • Temperature • Speed • Duration • Automated decapping • Specimen separated into aliquots • Aliquot labeling • In Vivo Haemolysis due to patient condition • Proper labeling • Mode of collection • Needle gauge • Position of arm • Location of venipuncture • Antiseptic used • Tourniquet time • Syringe transfer • Specimen tube • Type • Under/over filled • Order of draw • Under/over mixing • Origin of specimen • Vertical or horizontal transportation of tube • Pneumatic tube transport • Courier transport • Transportation duration • Centrifugation prior to transfer • Transfer temperature

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