Factors affecting assay of myoglobin by complement fixation or immunodiffusion.

Quantitative micro-scale complement-fixation and semi-quantitative gel immunodiffusion assays for myoglobin have been evaluated experimentally and used to measure myoglobin in the sera of more than 1500 patients. We report certain problems, observations, and methodological improvements. Serum caused enhancement of precipitin lines in the immunodiffusion assay. In the complement-fixation assay, serum interfered both by enhancing or inhibiting complement fixation. Suggested modifications in the complement-fixation assay are: (a) routine threefold or greater dilution of serum with buffer and (b) use of serum-based rather than buffer-based calibration materials in preparing standard curves. Three cycles of freeze-thaw of serum or buffer decreased detectable myoglobin by 50-100%. We conclude that certain serum-related factors and freeze-thaw phenomena are important causes of error in immunological assays for myoglobin.