Dilution of plasma with Tris buffer increases measured catecholamines in plasma

Area of Science:

• Biochemistry
• Analytical Chemistry
• Clinical Chemistry

Background:

• Radioenzymatic assays are commonly used for measuring catecholamines in plasma.
• Inhibition of methylation is a known issue in these assays, affecting accuracy.
• Plasma sample dilution is often employed to mitigate assay interferences.

Purpose of the Study:

• To investigate the impact of plasma sample dilution on catecholamine concentration measurements.
• To assess the effect of Tris buffer dilution on analytical recovery and radioactivity in catecholamine assays.
• To explore the potential mechanisms behind observed changes in catecholamine measurements upon dilution.

Main Methods:

• Human plasma samples were diluted 10-, 50-, and 100-fold using Tris buffer (100 mmol/L, pH 8.6).
• Analytical recovery of internal standards was evaluated.
• Radioactivity counts were measured to assess changes in catecholamine concentration.
• The influence of Tris buffer on catecholamine and conjugated catecholamine binding was examined.

Main Results:

• Plasma dilution with Tris buffer improved analytical recovery of internal standards.
• Dilution did not result in a proportional decrease in counted radioactivity.
• An unaccounted-for increase in radioactivity led to a significant rise in calculated catecholamine concentrations.
• These findings suggest Tris buffer may release catecholamines bound to plasma components.

Conclusions:

• Tris buffer dilution of plasma samples can improve analytical recovery in catecholamine assays.
• An observed increase in radioactivity suggests Tris buffer disrupts catecholamine binding to plasma components.
• Further research is needed to identify the low-molecular-mass component responsible for catecholamine binding.