Much has been published on the topic of method validation but a consensus protocol on how to perform the task is yet to be found.
This could be partly due to the fact that different analytical technologies have different requirements on which validation parameters that need to be addressed or that local initiatives by national societies in the clinical chemistry field were not discussed and spread at international level (4).
For example, carryover should be investigated in a chromatography-based method while it is not applicable in an ELISA.
The aim of the present work was to present straightforward step-by-step standard operating procedures (SOPs) for the validation of methods in which an analyte is determined in a biofluid matrix; the SOPs have been developed with the intention that they should be possible to follow without any advanced prior training.
Standard operating procedures for 10 different validation parameters are presented.
If a method is developed in-house, a full validation should be performed, meaning that all parameters should be investigated.
Last, these CSF biomarkers are applied in clinical studies on disease pathogenesis, and many research reports present novel biomarker candidates.
The vast majority of such fluid biomarkers are low-abundance proteins, for which antibody-based immunoassays, often in the enzyme-linked immunosorbent assay (ELISA) format, is needed to get enough analytical sensitivity.
This last item is sometimes overlooked; the intended use of a method needs to be carefully specified before any time consuming and costly validation experiments are performed. However, this paper will now focus on the validation of methods used to determine analyte concentrations in biofluids.
The aim of a method validation is to present objective evidence that a method fulfills the requirements for its intended use.
Although much has been published on which parameters to investigate in a method validation, less is available on a detailed level on how to perform the corresponding experiments.
The intended use for such a method could be to use the outcome as a diagnostic marker and in this case some evidence should be in place showing that there is a disease-dependent change in the analyte concentration in a biological sample.
Furthermore, the magnitude of the change should have an impact on the acceptable variability of the method, i.e., if the change is small the higher is the demand on the precision and on the analytical sensitivity and specificity.