A three-point identity criteria tool for establishing product identity using icIEF method

Area of Science:

• Biopharmaceutical analysis
• Protein characterization
• Drug release testing

Background:

• Ensuring product identity is crucial for biopharmaceutical release testing to prevent drug misidentification.
• Monoclonal antibodies (mAbs) and related products require robust identity testing methods.
• Isoelectric focusing (IEF) is a common technique, but its application to mAb identity needs careful evaluation.

Purpose of the Study:

• To investigate the challenges in establishing a product identity method for monoclonal antibodies (mAbs) using isoelectric focusing (icIEF).
• To evaluate the effectiveness of different sample preparation approaches (native, enzymatic digestion, reduction) combined with icIEF for mAb identity.
• To assess the suitability of icIEF as a stand-alone method for monoclonal antibody identity confirmation.

Main Methods:

• Utilized isoelectric focusing (IEF) coupled with capillary electrophoresis (CE) for analyzing monoclonal antibodies.
• Applied a three-point identity criteria tool: visual comparison, pI of individual peaks, and delta pIs (ΔpIs).
• Compared three sample preparation methods: native icIEF, enzymatic digestion followed by icIEF, and reduction followed by icIEF.

Main Results:

• The reduction approach followed by icIEF demonstrated higher potential for establishing the identity of mAb1 compared to native and enzymatic digestion methods.
• The three-point identity criteria tool aided in distinguishing mAb1 from other in-house mAbs.
• Isoelectric focusing (icIEF) generally lacks the required specificity for unequivocal identity establishment of monoclonal antibodies.

Conclusions:

• While icIEF can provide supporting data, it is not sufficiently specific as a stand-alone method for monoclonal antibody identity confirmation.
• A reduction approach prior to icIEF shows promise for enhancing identity assessment of mAbs.
• Risk analysis is recommended before implementing icIEF as the sole method for monoclonal antibody identity testing.