Platelet Aggregation After Gel Filtration

Ammit and O Neil (1991) used a quantitative bioassay of platelet aggregation for rapid and selective measurement of platelet-activating factor. 

Lumley P, Humphrey PPA (1981) A method for quantitating platelet aggregation and analyzing drug-receptor interactions on platelets in whole blood in vitro. J Pharmacol Meth 6 153-166 

Marguerie GA, Plow EF, Edgington TS (1979) Human platelets possess an inducible and saturable receptor specific for fibrinogen. J Biol Chem 254 5357-5363 Marguerie GA, Edgington TS, Plow EF (1980) Interaction of fibrinogen with its platelet receptor as part of a multistep reaction in ADP-induced platelet aggregation. J Biol Chem 255 154-161 

The entire procedure is performed in plastic (polystyrene) tubes at room temperature according to Marguerie et al. (1979). 

Blood is drawn from healthy adult volunteers, who had no medication for the last two weeks. Venous blood (8.4 ml) is collected into 1.4 ml ACD-solution and centrifuged for 10 min at 120 x g. The platelet-rich plasma (PRP) is carefully removed, the pH adjusted to 6.5 with ACD-solution and centrifuged at 285 x g for 20 min. The resulting pellet is resuspended in Tyrode s buffer (approx. 500 xl buffer/10 ml PRP). The platelet suspension is applied immediately to a Sepharose CL 2B column equilibration and elution at 2 ml/min flow rate is done with Tyrode s buffer without hirudin and apyrase. Platelets are recovered in the void volume. Final platelet suspension is adjusted to 4 x 108/ml. Gel-filtered platelets (GFP) are kept at room temperature for 1 h until the test is started. 

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