Technical methods. A simple method for the isolation of blood platelets.

Investigations of the physiological and biochemical properties of blood platelets in vitro have been severely hindered due to the lack ofasuitable method for their isolation.The platelets are fragile, they can be easily stimulated by a wide variety of agents leading to the release reaction and aggregation, also there are several plasma proteins loosely associated with the membrane.An ideal method for isolation should remove all foreign materials which are not an integral part of the platelet without causing any damage.Unfortunately, the conventional washing procedure causes gross and uncontrolled alteration of the platelets leading to variable results.The objective of this communication is to present a simple, economical, and reproducible method for isolating the platelets.This method is based on density gradient separation of platelets using sodium metrizoate as a cushion.Sodium metrizoate (Hypaque, Nyegaard Co, Oslo, Norway) combined with Ficoll is extensively used for the density gradient isolation of lymphocytes (Boldt, Skinner, and Kornfeld, 1972; Lopes, Nachbar, Zucker-Franklin, and Silber, 1973).Siliconized glass or plastic materials were used throughout this study.Normal blood from volunteer donors was collected in acid-citrate-dextrose (NIH formula A) in plastic bags and the red cells were removed by differential centrifugation.The plateletrich plasma (PRP) was recovered in a plastic bag in less than one hour after collection.Sodium metrizoate (32-8 %), a clear, sterile liquid with a density of 1 2, is supplied in glass ampoules (Nyegaard Co, Oslo, Norway).This material was diluted with distilled water to a density of 11.In a typical experiment, about 35 ml of PRP was slowly layered on top of 10 ml of diluted metrizoate in a 50-ml polycarbonate centrifuge tube.The inter- face between the PRP and metrizoate was gently disturbed with a siliconized stirring rod made from a pasteur pipette to produce a density gradient approximately 1 cm wide (fig la).The tubes were