What is the best method of preparing tissue for enzyme demonstration to preserve enzymatic activity?

• A. Unfixed frozen sections
• B. Formalin-fixed paraffin-embedded sections
• C. Alcohol-fixed frozen sections
• D. Fresh-fixed paraffin sections

Answer: (A)

Preserving enzymatic activity requires avoiding fixatives that cross-link proteins and inactivate enzymes. Chemical fixatives like formalin create cross-links that block enzyme active sites, and embedding in paraffin involves dehydration and heating that can destroy enzyme function. Freezing tissue without fixation keeps enzymes in their native state and preserves their active sites, allowing substrates to access and reveal activity during histochemical demonstrations. This unfixed frozen approach—rapidly freezing the tissue, sectioning in a cryostat, and applying the enzyme substrate—provides the best chance to visualize true enzymatic activity in situ. While other methods preserve morphology, they severely reduce or abolish enzyme activity, making them unsuitable for demonstrations of enzymatic function.