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The preparation and isolation of ¹⁴C-labelled egg white lysozyme and its utilization by in vitro cultured macrophage Blommers, Hendrik Willem


A preparative electrophoretic method was developed to isolate egg white lysozyme. The electrophoretic unit consisted of a slightly modified glass funnel in which a slab of polyacrylamide gel was suspended. The A₂₈₀ absorbing material extracted from the polyacrylamide gel travelled as a single staining band on quantitative polyacrylamide disc gels. Up to 25 mg. of apparently pure lysozyme was prepared by this method. Radioactive lysozyme was prepared by injecting young laying hens with a ¹⁴C ammo acid hydrolysate. By injecting the hydrolysate 5 hrs. postoviposition, lysozyme with a specific activity of 2 .6 x 10¹⁰ dpm/mole was prepared. Injection of the radioactive amino acids at intervals less than 5 hrs. postoviposition resulted in lower activity lysozyme. A theory has been suggested which could explain the generation of antibody diversity. In this theory (25, 26), the RNA portion of the RNA—antigen complex formed during an immune response, acts in an informational capacity. In order to investigate one aspect of the formation of these complexes, a radioactive antigen was required. The radioactive lysozyme isolated as described above was used. Nucleic acid extractions of macrophage incubated with radioactive lysozyme found little activity in the nucleic acid fractions. A thorough discussion in light of these results is presented along with some suggestions for further work.

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