Abstract
Summary
Purified myelin was prepared in high yield by a single centrifugation in the L-4 zonal centrifuge. A whole rabbit brain (8 g) homogenate in 7% sucrose was layered over a 10–35% sucrose gradient in the B-IV rotor and sedimented for 2 hr at 40,000 rpm. The gradient was removed by pumping 50% sucrose into the rotor with the recovery of 120 fractions of 13 ml each. Every fifth to tenth fraction was examined under the electron microscope and was assayed for RNA, DNA, sialic acids, and 10 enzymes. The latter included cytochrome oxidase (CyOx) to monitor for mitochondria, K-activated ATPase (K∗ATP) and acetylcholinesterase (AChE) for synaptic membranes, leucylglycinase for soluble and microsomes, acid phosphatase for lysosomes, and glucose-yields in fractions 90–110, in keeping with the presence of all subcellular constituents in these fractions.
We gratefully acknowledge the support of this project through NIH, Grant NB 08278.
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