Protein interferes with ionised calcium measurement at the reference electrode liquid junction

Area of Science:

• Clinical Chemistry
• Biophysical Chemistry

Background:

• Accurate measurement of ionized calcium (iCa) is critical in clinical diagnostics.
• Previous studies reported a positive correlation between serum protein and iCa using specific analyzers.
• The underlying mechanism for this observed correlation remained unclear, with potential contributions from Donnan effects or electrode artifacts.

Purpose of the Study:

• To investigate the cause of the reported positive relationship between protein concentration and ionized calcium.
• To differentiate between a true Donnan effect and an artifact related to reference electrode liquid junction potentials.
• To assess the impact of reference electrode junction solution tonicity on iCa measurements.

Main Methods:

• Ultrafiltration of human control serum to create sequential retentates with varying protein concentrations.
• Measurement of ionized calcium in retentates and filtrates using Nova 2 and Radiometer ICA1 analyzers.
• Systematic variation of reference electrode liquid junction solutions from hypertonic to isotonic.

Main Results:

• Ionized calcium increased with protein concentration using standard reference electrodes.
• This positive correlation reversed to a slight decrease with increasing protein when isotonic junction solutions were employed.
• Measurements with isotonic solutions paralleled those in the filtrates, suggesting artifactual interference.

Conclusions:

• The observed positive correlation between protein and ionized calcium is primarily an artifact of the reference electrode liquid junction potential.
• Protein's effect on the liquid junction potential, not a Donnan effect, explains the discrepancy in measurements.
• Careful consideration of reference electrode properties is essential for accurate ionized calcium determination in protein-rich samples.