BCDA

BCDA (5-bromo-4-chloroindoxyl acetate) is a chromogenic substrate of esterase used to potently detect the activity of esterase^[1]. In Vitro:The reference procedures for using BCDA as a chromogenic substrate for esterase in histochemistry are as follows:
1. Tissue Processing: Rat tissues are fixed with a 4% formaldehyde + 1% calcium chloride fixative at 0-2°C for 24-48 hours, and then transferred to a 0.88 M sucrose solution (containing 9 g/L gum acacia) for 24-48 hours of osmotic treatment.
2. Preparation of Staining Solution: Mix 2 mL of 0.1 M Tris buffer (pH 8.5), 1 mL of oxidant solution (50 mM potassium ferricyanide and 50 mM potassium ferrocyanide each), 5 mL of 2 M sodium chloride solution, and 2 mL of distilled water. Quickly add the substrate ethanol solution to prepare a 0.5 mM staining solution.
3. Tissue Sectioning and Staining: Frozen sections of kidney/liver are cut at a thickness of 2-5 μm, and intercostal muscle at 15-25 μm to preserve cellular structures. The sections are transferred to the staining solution and incubated at 37°C for 5-60 minutes (prolonged incubation should be avoided to prevent diffuse dye deposition).
4. Mounting and Observation: After neutralization with a 30% ethanol-0.1% acetic acid solution, the sections are mounted with glycerol gel or counterstained, and the amorphous dye deposition at esterase-active sites can be observed under a microscope.