ASTM D4012 - 81(2009)

    Standard Test Method for Adenosine Triphosphate (ATP) Content of Microorganisms in Water

    Active Standard ASTM D4012 | Developed by Subcommittee: D19.24

    Book of Standards Volume: 11.02


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    Significance and Use

    A rapid and routine procedure for determining biomass of the living microorganisms in cultures, waters, wastewaters, and in plankton and periphyton samples taken from surface waters is frequently of vital importance. However, classical techniques such as direct microscope counts, turbidity, organic chemical analyses, cell tagging, and plate counts are expensive, time-consuming, or tend to underestimate total numbers. In addition, some of these methods do not distinguish between living and nonliving cells.

    The ATP firefly (luciferin-luciferase) method is a rapid, sensitive determination of viable microbial biomass. ATP is the primary energy donor for life processes, does not exist in association with nonliving detrital material, and the amount of ATP per unit of biomass (expressed in weight) is relatively constant. (ATP per cell varies with species and physiological state of the organism.)

    This test method can be used to:

    Estimate viable microbial biomass in cultures, waters, and wastewaters.

    Estimate the amount of total viable biomass in plankton and periphyton samples.

    Estimate the number of viable cells in a unispecies culture if the ATP content (or if the average amount of ATP) per cell is known.

    Estimate and differentiate between zooplanktonic, phytoplanktonic, bacterial, and fungal ATP through size fractionation of water, and wastewater samples.

    Measure the mortality rate of microorganisms in toxicity tests in entrainment studies, and in other situations where populations or assemblages of microorganisms are placed under stress.

    1. Scope

    1.1 This test method covers the measurement of adenosine triphosphate (ATP) in microorganisms in concentrations normally found in laboratory cultures, waters, wastewaters, and in plankton and periphyton samples from waters.

    1.2 Knowledge of the concentration of ATP can be related to viable biomass or metabolic activity, or by utilizing an average concentration (or amount) of ATP per cell, an estimated count of microorganisms can be obtained in the case of unispecies cultures.

    1.3 This test method offers a high degree of sensitivity, rapidity, accuracy, and reproducibility. However, extreme care must be taken at each step in the analysis to ensure meaningful and reliable results.

    1.4 The analyst should be aware that the precision statement pertains only to determinations in reagent water and not necessarily in the matrix being tested.

    1.5 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.

    1.6 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.


    2. Referenced Documents (purchase separately) The documents listed below are referenced within the subject standard but are not provided as part of the standard.

    ASTM Standards

    D1129 Terminology Relating to Water

    D1193 Specification for Reagent Water


    ICS Code

    ICS Number Code 13.060.50 (Examination of water for chemical substances)

    UNSPSC Code

    UNSPSC Code


    DOI: 10.1520/D4012-81R09

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