Cytochemical Studies of the Action of Trypsin

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Abstract
Fixed and unfixed salivary glands of Drosophila melanogaster were treated with salt-free crystallized trypsin in a concn. of 0.1 mg./ml. dissolved in 0.05 M acetate or phosphate buffer at various pH levels, or in aqueous solns. adjusted for pH by adding traces of NaOH. Chromosomes of cells transferred directly from physiological salt soln. into buffered solns. of trypsin (pH 5.0-9.0) for 4 hrs. showed no effect of the enzyme treatment, although in many glands the intercellular materials were removed. Chromosomes in intact glands or smears fixed in acetic acid were attacked by trypsin in varying degrees, depending on the pH, solvent used, and the duration of treatment. Aqueous solns. of trypsin at pH 6.0 did not appreciably affect the chromosome structure within 24 hrs., whereas a treatment series of trypsin-water-.05 M phosphate buffer caused structural deformation. The course of such deformation included lateral swelling of the chromosomes, followed by the appearance of scattered and discrete chromomeres. Since treatment of intact glands or chromosome smears with desoxyribonuclease before and after trypsin treatment prevented deformation of chromosomes, these degradational changes are attributed to swelling in the presence of electrolytes of nucleic acids or nucleoproteins as a result of the hydrolysis, but not the dissolution, of chromosomal proteins. The need for using purified and carefully assayed samples of enzymes is stressed, if valid interpretations of chromosomal structure and cellular organization are to be obtained.

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