Changing Method Protocol

It is possible to switch kit reagents or change assay conditions if this suits the specific requirements of the study. Assay conditions, for example, incubation times, can be changed from the method supplied in the kit protocol to suit the drug development program. 

Incubation time, if shortened, may have an effect on the assay range and may make the assay more likely to suffer from drift. Incubation times, if increased, may help with assay drift and improve the assay range. However, increasing the overall assay time can decrease the potential sample throughput. Incubation temperatures may affect the assay range and the assays robustness. 

DeSilva,B., Smith, W., Weiner, R., Kelley, M., Smolec, J.,Lee,B.,Khan,M.,Tacey,R.,Hill, H., and Celniker, A. (2003) Recommendations for the bioanalytical method validation of ligand binding assays to support pharmacokinetic assessments of macromolecules. Phar maceutical Research, 20, 1885 1900. 

Passing, H., and Bablok, W. (1983) A new biometrical procedure for testing the equality of measurements from two different analytical methods. Application of linear regression procedures for method comparison studies in clinical chemistry. Part I. Journal of Clinical Chemistry and Clinical Biochemistry, 21, 709 720. 

Manley, S.E., Stratton, I.M., Andrew, H Neil, W Ross, I.S., Holman, R.R., Turner, R.C., and Matthews, D.R. (1997) Approach to maintaining comparability of biochemical data during long term clinical trials. Clinical Chemistry, 43, 1913 1918. 

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Method validation is closely related to method development. When a new method is being developed, some parameters are being evaluated already during the development stage while in fact this forms a part of the validation stage [15]. On the other hand, a validation study may indicate that a change in the method protocol is necessary, which may then require revalidation [58]. 

Robustness and ruggedness experiments are used to demonstrate how reproducible a method is when conditions vary. An assay method protocol defines the exact steps to be followed to ensure an accurate and precise result. Unfortunately, slight-to-large deviations occur in the everyday process, which may or may not impact the result. Assay conditions like incubation temperature or exposure to light define the robustness of the assay, whereas changes to the routine, for instance, multiple analysts or different instruments, define the ruggedness. The actual batch size for a routine sample analysis run is frequently overlooked but is often an impactful ruggedness measurement that should be assessed. 

It is a common mistake to attempt to combine method development with validation. Not only are the approaches to these two phases fundamentally different, making significant changes to method protocols during an SLV is likely to invalidate earlier data and result in more work being required. 

BERM-5 presented the changing outlook of the AOAC International in recognizing the usefulness of incorporating RMs for use in conjunction with their methods validation protocol. 

After the integration of this synthetic method into a fully compatible protocol, the positions for electronic changes with various substituents were rationally predicted through the deliberation of computational studies the Ri position of the... 

For systems that do not offer online HIAR, these procedures must be performed manually or off-line prior to loading slides on the instrument. Online HIAR methods are usually found as part of the closed-type automated staining systems, and are therefore less flexible in terms of what a technician can do to change the HIAR portion of a staining protocol to help optimize staining for particularly difficult antibodies. In spite of this, the ability to perform online HIAR is advantageous for many antibodies because it is extremely consistent and frees up technician time to complete other laboratory tasks. 

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