Preparation of Human Erythrocyte Ghosts in Isotonic Solution: Haemoglobin Content and Polypeptide Composition

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Abstract
Human red blood cell ghosts were prepared by dielectric breakdown of the cell membrane and subsequent electrical hemolysis in isotonic and isoionic solutions. Almost complete removal of Hb (to about 0.1%) was achieved by subjecting the erythrocytes suspended in isotonic solution to 3 consecutive electric field pulses of 16 kV/cm in the presence of 10 mM EDTA, followed by dilution with EDTA-free isoionic medium. The cup-shaped ghost cells were electrically homogeneous. The polypeptide composition of the electrically prepared ghost cells, as analyzed by polyacrylamide electrophoresis in 1% SDS [sodium dodecyl sulfate], revealed a pattern similar to ghost cells prepared by osmotic hemolysis. The band 6, corresponding to glyceraldehyde-3-phosphate-dehydrogenase protomer, was absent from the electrically prepared ghost cells. Also the band 8, which is eluted at low ionic strength, appeared to be very prominent in the electrical preparations.