Isolation of Plasma Membrane from Porcine Spermatozoa

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Abstract
A procedure is reported for isolating plasma membrane from porcine spermatozoa. Sperm were separated from gel particles and seminal plasma and subjected to low-intensity sonication that fragmented the plasma membrane, but minimized damage to the acrosome, nucleus and mitochondria. After cytoplasmic droplets and plasma membrane-depleted sperm were removed by differential centrifugation, the plasma membrane fragments were purified by flotation through a discontinuous sucrose gradient. During isolation, the effectiveness of each step was evaluated by light or electron microscopy. Plasma membrane fragments were identified using a stain specific for that membrane. Preparations were shown, by morphometry, to contain at least 70 per cent plasma membrane.