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Metabolism and Enzymology

Histochemical Technique: Densitometry of Nanogram Quantities of Proteins Separated in One-Dimensional Microslab Gels ¹

Department of Biological Science, Unit I, Florida State University, Tallahassee, Florida 32306-3050

We describe methods for densitometry of electrophoretically separated proteins in 25-millimeter microslab gels. The methods are sufficiently sensitive to use with several individually excised plant cells, for which we describe an extraction procedure. In brief, submicrogram samples are excised from freeze-dried plant tissue. Extraction takes place under oil in microliter droplets of detergent. Proteins are separated by one-dimensional microelectrophoresis (HM Poehling, V Neuhoff 1980 Electrophoresis 1: 90-102) and then stained by a sensitive Coomassie procedure (V Neuhoff, R Stamm, E Hansjorg 1985 Electrophoresis 6: 427-448). The resulting profile is scanned by a computerized densitometer based on the Leitz Diavert MPV Microphotometer. Evaluations and typical data demonstrate the high performance of this system.