Introduction

0.1 The ISO 16140 series

The ISO 16140 series has been expanded in response to the need for various ways to validate or verify test methods. It is the successor to ISO 16140:2003. The ISO 16140 series consists of six parts with the general title, Microbiology of the food chain — Method validation:

— Part 1: Vocabulary;
— Part 2: Protocol for the validation of alternative (proprietary) methods against a reference method;
— Part 3: Protocol for the verification of validated reference methods and validated alternative methods in a single laboratory;
— Part 4: Protocol for method validation in a single laboratory;
— Part 5: Protocol for factorial interlaboratory validation for non-proprietary methods;
— Part 6: Protocol for the validation of alternative (proprietary) methods for microbiological confirmation and typing procedures.

ISO 17468 is a closely linked International Standard, which establishes technical rules for the development and validation of standardized methods.

In general, two stages are needed before a method can be used in a laboratory.

— The first stage is the validation of the method. Validation is conducted using a study in a single laboratory followed by an interlaboratory study (see ISO 16140‑2, ISO 16140‑5 and as described in this document). In the case when a method is validated within one laboratory (see ISO 16140‑4), no interlaboratory study is conducted.
— The second stage is method verification, where a laboratory demonstrates that it can satisfactorily perform a validated method. This is described in ISO 16140‑3. Verification is only applicable to methods that have been validated using an interlaboratory study.

In general, two types of methods are distinguished: reference methods and alternative methods.

A reference method is defined in ISO 16140‑1:2016, 2.59, as an “internationally recognized and widely accepted method”. The note to entry clarifies that “these are ISO standards and standards jointly published by ISO and CEN or other regional/national standards of equivalent standing”.

In the ISO 16140 series, reference methods include standardized reference (ISO and CEN) methods as defined in ISO 17468:2016, 3.5, as a “reference method described in a standard”.

An alternative method (method submitted for validation) is defined in ISO 16140‑1:2016, 2.4, as a “method of analysis that detects or quantifies, for a given category of products, the same analyte as is detected or quantified using the corresponding reference method”. The note to entry clarifies that: “The method can be proprietary. The term ‘alternative’ is used to refer to the entire ‘test procedure and reaction system’. This term includes all ingredients, whether material or otherwise, required for implementing the method.”.

ISO 16140‑4 addresses validation within a single laboratory. The results are therefore only valid for the laboratory that conducted the study. In this case, verification (as described in ISO 16140‑3) is not applicable. ISO 16140‑5 describes protocols for non-proprietary methods where a more rapid validation is required or when the method to be validated is highly specialized and the number of participating laboratories required by ISO 16140‑2 cannot be reached. ISO 16140‑4 and ISO 16140‑5 can be used for validation against a reference method. ISO 16140‑4 (qualitative and quantitative) and ISO 16140‑5 (quantitative only) can also be used for validation without a reference method.

The flow chart in Figure 1 gives an overview of the links between the different parts mentioned above. It also guides the user in selecting the right part of the ISO 16140 series, taking into account the purpose of the study and the remarks given above.

Figure 1 — Flow chart for application of the ISO 16140 series

NOTE In this document, the words “category”, “type” and/or “item” are sometimes combined with “(food)” to improve readability. However, the word “food” is interchangeable with “feed” and other areas of the food chain as mentioned in Clause 1.

This document, ISO 16140-6, is somewhat different from the other parts in the ISO 16140 series in that it relates to a very specific situation where only the confirmation procedure of a method is to be validated [e.g. the biochemical confirmation of Enterobacteriaceae (see ISO 21528‑2)]. The confirmation procedure advances a suspected (presumptive) result to a confirmed positive result. The validation of alternative typing techniques (e.g. serotyping of Salmonella) is also covered by this document. The validation study in this document clearly defines the selective agar(s) from which strains can be confirmed using the alternative confirmation method. If successfully validated, the alternative confirmation method can only be used if strains are recovered on an agar that was used and shown to be acceptable within the validation study. Figure 2 shows the possibilities where an alternative confirmation method validated in accordance with this document can be applied (see text in the boxes).

Figure 2 — Use of validated alternative confirmation methods (described in this document)

EXAMPLE

An example application of a validated alternative confirmation method is as follows.

An alternative confirmation method based on ELISA has been validated to replace the biochemical confirmation for Salmonella as described in ISO 6579‑1. In the validation study, XLD (mandatory agar in accordance with ISO 6579‑1) plus BGA and a specified chromogenic agar (two optional agars for second plating in accordance with ISO 6579‑1) were used as the agars to start the confirmation. The validated confirmation method can be used to replace the biochemical confirmation under the following conditions:

— by laboratories using the ISO 6579‑1; or
— by laboratories using an ISO 16140‑2 validated alternative method that refers to ISO 6579‑1 for confirmation; or
— by laboratories using an ISO 16140‑2 validated alternative method that starts the confirmation from XLD and/or BGA agar and/or the specified chromogenic agar.

The validated confirmation method cannot be used under the following conditions:

— by laboratories using an ISO 16140‑2 validated alternative method that refers only to agars other than those included in the validation to start the confirmation (e.g. Hektoen agar and SS agar only); or
— by laboratories using an ISO 16140‑2 validated alternative method that refers only to a confirmation procedure that does not require isolation on agar.

0.2 Validation and verification of methods for the microbiological confirmation and typing procedures

The procedure described in this document is intended for the “full” validation of alternative (proprietary) methods for microbiological confirmation and/or typing, hereafter referred to as “alternative confirmation methods”.

During the validation study, the performance of the alternative confirmation method is compared to the performance of the reference confirmation procedure.

The procedure for verification of alternative confirmation methods in a single laboratory is described in ISO 16140‑3.

BS EN ISO 16140-6:2019

Microbiology of the food chain. Method validation - Protocol for the validation of alternative (proprietary) methods for microbiological confirmation and typing procedures

Current

Published:

31 Dec 2019