Rapid whole-blood platelet counting using an electronic particle counter

A Coulter electronic particle counter, model A with an orifice tube for the counter, aperture 70,u, was used, and saline, 0-85 g.NaCl per 100 ml.distilled water, filtered to remove most particles (M.P.C. stainless steel laboratory filter, grade No. 100)1.Flat-bottomed siliconed glass containers, 25 mm.internal diameter, are required.METHOD Blood is collected into E.D.T.A. (4 mg. of ethylene diamine tetra-acetic acid, dipotassium salt per 2-5 ml.blood, dried on the surface of the container).Blood, 0-2 ml., is added to 2-3 ml.saline in a siliconed glass con- tainer and mixed.Centrifuge for 20 seconds at 750 to 1,250 r.p.m.Within this range the speed is not critical, but the duration of spinning is critical.After less than 20 seconds there is too high a red cell count in the super- natant fluid, and after 30 seconds there is loss of platelets into the deposit, the larger platelets being lost first.Each centrifuge needs testing at various speeds within the range indicated.Rapid acceleration to the required speed is essential, and was obtained on the model used by temporarily switching to a higher stop for 10 seconds.Smooth slowing is also required.The particular centri- fuge used when switched off at 1,000 r.p.m. ceased spinning about 40 seconds later.(Its radius was 14 cm., and the centrifugal force is proportional to the square of the radius.)Carefully pipette off the supernatant fluid.Add 0-5 ml. of supernatant to 99 5 ml.filtered saline and mix.This diluted sample is counted in the counter at settings advised for each machine by Coulter Electronics Ltd.This particular machine and orifice tube, set at threshold 15, is used at aperture current setting 7 (A.C.S. 7) to count all particles 3-6pu' and above and at A.C.S. setting at 3 to count all particles 38-3j.3 and above.