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Highly purified preparations of poliomyelitis virus, strain MEF-1, have been shown by analytical ultracentrifugation to contain only one homogeneous component.1 Electron microscopic and other studies have revealed this component to consist of spherical particles approximately 27 mu in diameter which are readily aligned in two-dimensional, close-packed arrays and with which infectivity is associated.1' 2 The present report is concerned with the preparation and the description of properties of three-dimensional crystals, readily visible in the light microscope, which were obtained from a highly purified and concentrated preparation of such virus particles. Although several plant viruses have been crystallized' since the initial crystallization of tobacco mosaic virus by Stanley in 1935,4 this is believed to be the first report of the crystallization of a virus affecting man or animals. Crystallization.-The source of virus was the fluid harvested from monkey kidney tissue cultures infected with strain MEF-1 poliomyelitis virus and provided by the Connaught Laboratories, Toronto. The initial purification procedure, which has been previously outlined,' included methanol precipitation, butanol extraction, ultracentrifugation, nuclease digestion, and electrophoretic fractionation. Further purification of the preparation was achieved by sedimentation through a sucrose density gradient in a swinging cup rotor (Spinco SW-39), as described by Brakke for plant viruses.5 In this way approximately 1.2 mg. of nucleoprotein material was obtained as a single sedimenting band in 0.73 ml. of 0.14 M NaCl containing approximately 30 per cent sucrose. This represented the virus concentrated from 15 liters of infected monkey kidney tissue culture fluid. The preparation was diluted to 2.0 ml. with 0.14M NaCl and was sedimented by ultracentrifugation in an average centrifugal field 100,000 X g for 2.5 hours. The clear, gel-like pellet was covered with 0.3 ml. of unbuffered 0.14 M NaCl at pH 5.9 and gently rocked overnight at 40 C. After this treatment the pellet appeared white and opaque, and upon suspension discrete crystals were seen. After removing a small fraction of the suspended crystals for further observation, the mother-liquor was removed from the remaining crystals. The latter were washed twice with isotonic saline and the washings pooled with the mother-liquor. The crystals were readily dissolved by warming to room temperature in 0.14 M NaCl adjusted to pH 8.5 with dilute NaOH. The virus was recrystallized as follows: An aliquot of the dissolved crystals was diluted with a small volume of 0.14M NaCl buffered with 0.001M phosphate at pH 7.6, and the virus was then sedimented in a sealed capsule6 in the swinging cup rotor of the ultracentrifuge. After removal of the supernatant fluid, the pellet was suspended in a small volume of isotonic saline buffered with 0.01 M phosphate at pH 5.9 and allowed to stand at 4° C. Small crystals were seen by microscopic examination. After low-speed centrifugation, the mother-liquor was removed from the re-

CRYSTALLIZATION OF PURIFIED MEF-1 POLIOMYELITIS VIRUS PARTICLES