Which of the following methods depends on cleavage of synthetic substrates by an active serine protease?

• A. Chromogenic
• B. Photo-optical
• C. Mechanical
• D. Immunodiffusion

Answer: (A)

Chromogenic assays are built around a synthetic substrate linked to a color-producing group. When an active serine protease cleaves that substrate at its specific peptide bond, the chromophore is released or unmasked, yielding a color change that can be measured spectrophotometrically. The rate of color development directly reflects the enzyme’s activity, making this approach ideal for quantifying serine proteases like thrombin or factor Xa in hemostasis testing. Immunodiffusion relies on antibody-antigen diffusion and does not measure enzymatic cleavage; mechanical methods assess clot formation by physical changes rather than substrate cleavage; photo-optical methods may track optical changes from various processes but aren’t defined by cleavage of a synthetic substrate by an active serine protease.