HISTOCHEMICAL STUDIES IN HURLER'S DISEASE: A NEW METHOD FOR LOCALIZATION OF ACID MUCOPOLYSACCHARIDE, AND AN ANALYSIS OF LEAD ACETATE "FIXATION"

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Abstract
From these investigations, optimal preservation of the Hurler acid mucopolysaccharide can be achieved by cutting frozen sections of unfixed tissue, with subsequent fixation in a mixture of tetrahydrofuran-acetone (1:1) for at least 20-30 minutes. In the liver the acid mucopolysaccharide appears to be largely extracellular. It is seen either as homogeneous diffuse or as finely granular material localized to "islets." The relation between these 2 forms remains to be determined. In the dermis the acid mucopolysaccharide appears to be extracellular and situated between collagen fibers. The "clear spaces" seen in paraffin sections following the dissolution of acid mucopolysaccharide apparently represent foci of swelling of the acid mucopolysaccharide exposed to a solvent medium. The metachromasia of the Hurler acid mucopolysaccharide is best demonstrated following the above fixation, when the sections are stained in 0.5% toluidine blue in 25% acetone. The method described appears suitable for diagnostic skin or liver biopsies. It is possible to remove the crystalline deposit from sections of tissue fixed in lead acetate solution. Both an iodine-sodium thiosulfate sequence and disodium versenate remove the crystalline deposit entirely, but the latter also reduces the staining of tissues with toluidine blue. The lead acetate fixative preserves the Hurler acid mucopolysaccharide poorly and interferes with the meta-chromatic staining of the acid mucopolysaccharide with toluidine blue. Lead acetate competes with this dye in combining with the Hurler acid mucopolysaccharide, not only when the tissue is fixed in this substance, but also when other fixatives have been used, or when sections are exposed to a mixture of lead acetate and toluidine blue. The combination of lead acetate and Hurler acid mucopolysaccharide is reversible by prolonged exposure to toluidine blue solution, although it appears to be a stronger combination than that of lead acetate with either heparin or commercial chondroitin sulfate.

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