Luciferase preparation, assay conditions and calculated extraction efficiency as factors in the estimation of soil ATP content
Several factors affecting the measurement of soil adenosine 5′-triphosphate (ATP) using a crude (macerated firefly tail) preparation of luciferase and a luminometer were investigated. These factors were luciferase preparation purity (crude or purified), use of Tris(hydroxymethyl)methylamine (Tris) or sodium arsenate (arsenate) buffer for the luciferase preparation and the bioluminescent reaction, storage temperature of luciferase throughout the assay (20°C or 1°C), modes of measuring bioluminescence (peak height or integration of bioluminescence decay) and efficiency of extraction (recovery) of ATP from soil. Crude luciferase produced a linear relationship between bioluminescence and ATP concentration very similar to that produced by purified luciferase and could be stored at 20°C for 3 h without deterioration. Arsenate was, overall, the preferred buffer for the assay. Integrating light output within 15 s of mixing luciferase-luciferin and ATP avoided interference from other reactions in the crude extracts. The procedure used to calculate the ATP concentration and the amount of exogenous ATP added to soil both affected the calculated efficiency of ATP recovery. Thus an assay for soil ATP content using crude preparations has been developed which is as sensitive as those using purified preparations. © 1986 Springer-Verlag.
West, AW; Sparlings, GP; Cowlings, JC; Tatel, KR; Reynolds, J
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