A Demineralization Procedure for Enzymatic Histochemical Use: A Quantitative Succinic Dehydrogenase Assay

Thinnest practical slices of bones or teeth are suspended at 4 C in a 10% solution of EDTA in 0.1 M tris buffer brought to pH 6.95 with KOH pellets. The solution is stirred moderately and continuously with a magnetic stirrer until specimens are demineralized. Unwashed specimens, taken directly from the demineralizing fluid, are frozen on a block of CO₂ ice, mounted on a tissue carrier and sectioned at 6 μ in a cryostat. Histochemical stains conducted on sections stored in a slide holder enclosed in a plastic bag in a refrigerator for as long as 2 wk were successful. Quantitative studies on the preservation of succinic dehydrogenase showed nearly all of it present in specimens demineralized for 2 days, and approximately 50% remaining in specimens demineralized for 7 days. Qualitative studies with other dehydrogenases indicate that several may be similarly affected.     

The effect of temperature on the rate of conversion of p,p-DDT to p,p-DDE in brook trout (Salvelinus fontinalis).

Brook trout (Salvelinus fontinalis) were maintained in flowing fresh water at 2, 10, and 18 degrees C and injected intramuscularly with p,p-[-14C]DDT. Fish were killed at intervals up to 16 weeks after injection, and analyzed by a combination of thin-layer chromatography, gas-liquid chromatography, and liquid-scintillation counting. Only one labelled metabolite, p,p-DDE, was detected during the experimental period. Its rate of production varied with temperature; after 16 weeks, about 9, 13, and 19 percent of the original p,p-DDT in fish maintained at 2, 10, and 18 degrees C, respectively, had been converted.