Optimization of an enzyme-coupling method by spectrophotometer for serum adenosine deaminase: As a candidate...

Area of Science:

• Biochemistry
• Clinical Chemistry
• Enzyme Assays

Background:

• Adenosine deaminase (ADA) is crucial for adenosine metabolism.
• Existing ADA measurement kits show variable results, hindering accurate clinical interpretation.
• Standardization of ADA assays is needed for reliable diagnostic outcomes.

Purpose of the Study:

• To optimize and validate a reference measurement procedure (RMP) for serum ADA.
• To establish a standardized method for routine ADA testing.
• To improve the accuracy and comparability of ADA measurements across different laboratories.

Main Methods:

• Enzyme-linked assay using ADA, purine-nucleoside-phosphorylase, xanthine-oxidase, and peroxidase.
• Optimization of the assay using Response Surface Methodology.
• Validation of the RMP through inter-laboratory comparisons and assessment of precision, linearity, and interference.

Main Results:

• The optimized RMP demonstrated excellent intra-assay (0.44%) and inter-assay (≤1.01%) precision.
• High linearity was observed up to 351.9 U/L (R² = 0.9998) with minimal interference (<5%).
• Inter-laboratory reproducibility was high (R² ≥ 0.9975), and a reference interval of 11.7-38.5 U/L was established.

Conclusions:

• The developed candidate RMP for serum ADA offers a reliable method for standardizing routine clinical assays.
• Calibration of a routine system using the RMP significantly improved its accuracy (deviation reduced from -55.9% to -3.7%).
• This standardized procedure has the potential to enhance the diagnostic utility of ADA measurements in clinical settings.