The volume of fixing fluid should be

• A. 15-20 times the amount of specimen
• B. 5-10 times the amount of specimen
• C. 10-15 times the amount of specimen
• D. 20-30 times the amount of specimen

Answer: (A)

The important idea here is that fixation works by the fixative diffusing into the tissue, so having enough fixative relative to the tissue ensures the interior of the specimen is preserved just as well as the surface. If the volume of fixative is too small, the fixative near the tissue gets used up and deeper cells may not fix properly, leading to autolysis and poor morphology. Using about 15–20 times the tissue volume keeps a strong enough reservoir of fixative so diffusion can proceed uniformly throughout the specimen, maintaining the concentration gradient that drives penetration and preventing dilution or depletion effects. Ratios on the lower end (5–10x or 10–15x) risk underfixation for larger or denser tissues, while much larger volumes (20–30x) tend to waste fixative without significantly improving results. So, 15–20 times the tissue volume is the practical, effective choice.