ISO 11816-2:2024

Overview

ISO 11816-2:2024 - Milk and milk products - Determination of alkaline phosphatase activity - Part 2: Fluorimetric method for cheese specifies a fluorimetric assay to measure alkaline phosphatase (ALP) activity in cheese. The method is applicable to soft, semi‑hard and hard cheeses provided mould is limited to the surface (not for blue‑veined cheeses). Measurements use a Fluorophos® test system and an instrument (current FLM300 replacement for FLM200) capable of reading ALP activity in the supernatant up to 7 000 mU/kg.

Key topics and technical requirements

• Principle: Continuous fluorimetric direct kinetic assay. A non‑fluorescent monophosphoric‑ester substrate is hydrolysed by ALP to a fluorescent product; fluorescence kinetics are recorded at 38 °C over a 3‑minute run (first minute = equilibration; activity measured during minutes 2–3).
• Reagents & test system: Uses Fluorophos substrate and DEA buffer to prepare a working substrate. Prepared working substrate must pass an analogue‑to‑digital (A/D) control (readings ≤ 1 200 FLU).
• Apparatus & instrument: The Fluorophos instrument (FLM300) includes a heater block and upgraded electronics/user interface; instrument performance verification, daily tests and calibration procedures are defined.
• Sampling & sample prep: Specific sampling rules apply (Clause 7), with special conditions for large hard cheeses. Sample preparation and supernatant separation are covered before fluorimetric determination.
• Controls & calibration: Daily instrument checks, reagent controls, calibration routines and recommended negative/positive controls and interfering‑substance tests are described.
• Calculation & expression: Results expressed as milliunits of enzyme activity per kilogram (mU/kg). Clauses cover calibration ratio, calculation for supernatant and cheese, and reporting.
• Precision & validation: Repeatability and reproducibility data are provided from interlaboratory studies; annexes include examples and study details.

Applications and users

• Who uses it: Dairy testing laboratories, cheese manufacturers’ quality control teams, food safety regulators, accreditation bodies and instrument/manufacturers supplying fluorimetric ALP kits.
• Why use it: To verify pasteurization adequacy, monitor processing and storage, detect residual enzyme activity indicative of inadequate heat treatment or microbial activity, and to comply with regulatory/industry requirements for milk and cheese safety.

Related standards

• ISO 11816 series (other parts for milk and milk products) and national dairy testing standards. Users should refer to the full ISO 11816-2:2024 text for detailed procedures, sampling rules (Clause 7), controls and calculation formulas.