Study objective: Measurement of pleural fluid adenosine deaminase (ADA) levels is useful in the differential diagnosis of pleural effusions. However, at ambient temperatures, the levels of this enzyme decline with time. The purpose of the present study was to identify, test, and optimize additives that stabilize ADA, consequently eliminating the need for dry ice or other cold specimen transport media.
Design: A preliminary screen of historically proven stabilizing agents for specific proteins demonstrated effectiveness of glycerol for maintenance of pleural fluid ADA levels. Systematic studies for exploitation of the glycerol effect included the following: (1) supplements to the glycerol of promising alternate compounds, (2) long-term stability studies at ambient and elevated temperatures, (3) a field test of an effective mixture as a means for reduction of specimen transport costs, (4) thermal stability studies for optimization of the agents for use at otherwise denaturing temperatures, and (5) inclusion of pleural fluids from patients with a variety of etiologies, including tuberculous pleurisy, in order to gauge the effectiveness of the stabilizing agents on both the high and low molecular weight forms of ADA.
Results: A mixture of glycerol and ethylene glycol, each at 5% concentration, maintained pleural fluid ADA levels for at least 21 days at both room temperatures and 37°C. A field test of 32 pleural fluids found that ADA levels in specimens containing this mixture, sent to the laboratory by surface mail at ambient temperatures, were nearly identical to those in aliquots of the same fluids shipped over dry ice. The bias in the measurement was 0.49 IU/L, with a precision of 2.49 IU/L. The correlation coefficient between the two measurements was 0.97. Thermal stability studies found that tuberculous pleural fluids containing 10% glycerol and 0.10 mol/L sodium sulfate maintained constant ADA levels for at least 10 days at 45°C, an otherwise denaturing temperature for nonstabilized specimens.
Conclusion: The addition of stabilizing agents to pleural fluid specimens allows the transport of those specimens to distant laboratories at ambient temperatures without a decline in the ADA levels. Employment of those agents will decrease the cost of the test and facilitate its use in second- and third-world countries.