Assuring Accuracy, Preventing Fraud
The Critical Role of Validation in Bioanalytical Methods
June 2024 | Author by Myriam Gueye, Segment Marketing Manager at Sartorius LPS

Harmonizing Regulatory Guidelines
for Assay Validation
A Focus on Ligand-Based Assays Using Octet BLI Systems

In the constantly advancing fields of scientific research andpharmaceutical development, the precision and reliability ofbioanalytical methods are critical. Kinetic-based assays,
particularly those employing Octet® BLI systems, havebecome fundamental due to their ability to provide real-time,label-free analysis of biomolecular interactions. Thoroughvalidation of these assays is crucial, as it ensures data
generation that is both accurate and reliable. This level ofdata integrity is vital for informed decision-making in clinicaldiagnostics and drug development.
The validation process for bioanalytical methods begins
with the determination of the standard deviation of theblank, a critical step that lays the foundation for establishing
the Detection Limit (DL) and the Quantitation Limit (QL) ofthe assay. These parameters are vital for defining thesensitivity and precision of the assay, which in turn, influences
the reliability of the data obtained.
Another key aspect of assay validation is defining the assay
range, which is the span between the lower limit ofquantitation (LLOQ) and the upper limit of quantitation(ULOQ). Range can also be established for ligand binding
assays and spans the lowest and highest affinity constant(KD) values obtained during assay qualification studies.
The robustness and stability of the assay are equally criticalfactors. Robustness refers to the assay's resilience to minor,
deliberate variations in the testing procedure, while stability
indicates the assay's ability to produce consistent results overtime and across various conditions. These characteristics arerigorously evaluated during method development toconfirm that the assay is suitable for real-world applicationand that the data generated is consistently reliable.

2

The assay validation process transcends the objective ofadhering to regulatory standards; it is fundamentally about
confirming that the assays are suitable for their designatedpurpose. This necessitates adapting the validation process toeffectively address the specific requirements and challenges
associated with kinetic-based assays.
Furthermore, the validation includes the assessment of theanalyte’s linear response. Linear regression analysis isperformed to establish the correlation between analyte
concentration and signal response. Acceptance criteria are
set for signal-to-noise ratio, standard deviation of the linear
response, and slope, all of which are critical to ensure theprecision and accuracy of the assay.
Stability testing forms an integral part of the robustness
evaluation, examining the assay's performance over time andunder various storage and handling conditions. This testing
serves as evidence of the assay's durability and theconsistency of the data it produces.
Octet® BLI 21 CFR Part 11 Compliance
Ensuring precision and safeguarding against data
manipulation are essential aspects of drug development andquality assurance. The Octet® BLI system from Sartorius isengineered to meet these demands, providing a range oftools and services that support adherence to good practice(GxP) standards. The platform includes installation andoperational qualification kits, performance qualification kits,
software validation packages, and biosensor validation
services, all of which contribute to the integrity and reliabilityof bioanalytical assays.

Installation
Qualification(IQ)

Operational
Qualification(OQ)
Analytical
Instrument
Qualification

MaintenanceCompliance
Assurance (PMOQ)

A critical component for maintaining data accuracy andsecurity is the Octet® 21 CFR Part 11 Software package,
which aligns with the FDA's 21 CFR Part 11 regulations.
This software suite includes features such as an audit trail,electronic signatures, and robust electronic record-keeping,which are vital for deterring fraud and maintaining dataintegrity. Sartorius professionals handle the installation toensure it is implemented correctly.
The Octet® GxP server is instrumental in record compliance,
and access to the computer hosting this software is tightly
controlled to prevent unauthorized use. The platform's
reliability is evidenced by its use in critical quality control
processes such as lot release and in-process testing assays,
including quantitation, potency assays, impurities testing,and ligand binding assays. Regulatory bodies haveacknowledged the precision of the Octet® BLI platform, asevidenced by the inclusion of Octet® BLI data in the approvalsubmissions for several drugs.
FDA Reminds Medical Device Manufacturers to
Scrutinize Third-Party-Generated Data
The validation of assays extends beyond the pursuit ofscientific precision; it acts as a safeguard against the risk of
fraudulent and unreliable data. In February 2024, the U.S.
Food and Drug Administration (FDA) has underscored theimportance of this issue, alerting the scientific community to
the severe implications that can result from compromised
data integrity.

Performance
Qualification(PQ)

The FDA's advisory serves as a potent reminder forlaboratories to maintain the highest standards of integrity
and accuracy in their testing procedures. It is a stark warningthat the manipulation or falsification of data can lead tosevere consequences, such as endangering patient safety,
the invalidation of research, and the potential for legal andregulatory action.
To mitigate these risks, the validation process for kineticbased assays is designed to be comprehensive andtransparent. It incorporates stringent checks and balances,including the verification of raw data, the authentication ofresults, and the implementation of quality control measuresthroughout the entire lifecycle of the assay. These measures
are critical in ensuring that the data generated is not only
scientifically robust but also ethically obtained and reported.
The FDA's emphasis on data integrity reinforces the need fora culture of quality in laboratory environments. Such a cultureis built upon the principles of good laboratory practices,diligent documentation, and the continuous training ofpersonnel. It is a culture that prioritizes the reproducibility ofresults and the traceability of data, recognizing that thereliability of laboratory testing is fundamental to bothscientific advancement and public trust.

3Conclusion
FDA’s Final Rule on Laboratory-Developed Tests
The FDA has issued a final rule to regulate laboratorydeveloped tests (LDTs) as medical devices, aligning themwith the oversight applied to other in vitro diagnostics under
the Federal Food, Drug & Cosmetic Act. In the past, LDTs
were exempt from FDA review due to their limited use andperceived lower risk. However, given the expanded use ofLDTs and their associated higher risks, the FDA has
introduced a comprehensive 528-page rule. This rule isintended to gradually eliminate this previous exemption overa period of four years, requiring all diagnostics, including
LDTs, to meet the same regulatory standards.

The harmonization of regulatory guidelines for assay
validation, with a particular focus on kinetic-based assays
using Octet® BLI systems, is a critical endeavor in the context
of scientific research and pharmaceutical development. The
comprehensive validation process, which includes
establishing detection and quantitation limits, defining assay
range, and ensuring robustness and stability, is not just aregulatory requirement but a commitment to scientificintegrity and ethical standards. The FDA's recent actions,including the final rule on LDTs and the emphasis on data
integrity, serve as evidence to the agency's dedication topatient safety and the reliability of medical devices.
Laboratories are now called upon to foster a culture ofquality, where good laboratory practices, accurate recordkeeping, and continuous training are the norm. As theindustry moves forward, the meticulous validation of assays isindispensable for the advancement of science, theprotection of public health, and the preservation of trust in
medical research and diagnostics.

Publications and Sources
1. The Impact of IVD Regulations on Manufacturing 		 Publication | Sartorius
2. Octet® BLI 21 CFR Part 11 Compliance Checklist |		Sartorius
3. Fraudulent and Unreliable Laboratory Testing Data in		 Premarket Submissions: FDA Reminds Medical Device		 Manufacturers to Scrutinize Third-Party-Generated Data |		FDA4. Federal Register: Medical Devices; Laboratory		Developed Tests

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