TABLE 1.

Questions useful for identifying the information needed to define the measurand.

Question(s)	Answer and/or guidance for a specific product
What is the analyte? What is being detected? What is being counted?	The analyte is the entity measured by the analytical procedure. The analyte is culturable cells enumerated as colony forming units (CFU).
What is the matrix? Are there excipients? Stabilizers?	Matrix components are generally cryoprotectants, excipients, bulking agents for powder flow, etc.
Are there possible contaminants in the matrix?	Non-microbial contaminants: Carryover from fermentation media, leachables, and extractables from production systems. Microbial contaminants (both live and dead), remnants of cell-walls, cross-contaminants (from other strains produced in the same facility), and environmental contaminants.
Will the term “pure” be used to describe the ingredient?	Probiotic ingredients are often described as “pure” powders. The term “pure” is controversial but useful. The discipline of defining a measurand requires that the meaning of the controversial term, pure or purity, be defined if used. The probiotic ingredient (freeze dried cells) is usually a pure powder that may contain cryoprotectant and carryover of fermentation media. It does not contain excipients as do final formulated product.
Matrix: other components.	Is the probiotic ingredient a pure powder? Is the probiotic ingredient definition, above, used to describe the term pure? Is the probiotic ingredient in a solution or suspension? Include the solvent or suspension liquid in the measurand definition. Usually, there are no solvents in a freeze-dried product.
What is the decision unit (also known as parent body)? For what entity will the decision be made?	Options to consider for the decision unit: Laboratory sample, a batch of probiotic ingredient, a product lot. Composite sample or a single grab sample. Randomly selected from a bulk-capsule or finished capsule lot. The sample taken from the beginning, middle, or end of the batch, or at all three time points. The form of the sample is a bulk ingredient, formulated blend, capsules, sachets. R&D may conduct a study during process development to ensure the sample is representative, and that the uncertainty contribution from sampling is not of practical importance.
What is the physical form of the decision unit?	Powder, solution, etc. Describe the form.
Define the units for the quantity.	For example, the unit can be CFU/g or CFU/mL.
The information in the following two points is not needed to define the measurand; but is needed to complete the ATP. It is convenient to collect this information along with details for defining the measurand.
What is the concentration range of test results that should be reported by the analytical procedure?	The laboratory may extend that range to include concentrations for potentially OOS values. This information is usually provided by the development team.
What is the counting range?	There are different standards for the counting range. The counting range depends on the size of the Petri dish, the applied agar, the probiotic strain, etc. It is up to the manufacturer to assess the counting range and the linearity for a specific ingredient and/or product with the applied CFU method.

The answer for each question is either information applicable to the analytical probiotic enumeration procedure or guidance for a company’s specific product. These questions address both the probiotic ingredient and finished product.