Penicillan Binding Protein Assay青霉素与细胞膜蛋白结合实验
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Wash cells with 10 mM Tris pH 8
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French press
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Slow speed spin
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High speed spin
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Resuspend in 10 mM Tris
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Sonicate 2 x 15 sec to remove ß-lacatamases
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Wash in 10 mM Tris
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Resuspend in 10 mM Tris
- Store at ?70ºC until needed.
Assay:
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Incubate membranes with ~5 µCi 3 H-penicillin at 25ºC for 10 minutes
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Add 1000X cold penicillin
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SDS-PAGE
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Incubate in 1 M sodium salicylate for 30 minutes
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Dry at 80ºC for 2 hours under vacuum
- Expose to photographic film for 3-30 days.