Assay Packages: Cell Line Development

Assay families that satisfy testing needs include:

• Titer

  Productivity of the cell line to deliver sufficient quantities of your mAb influences its commercial potential. Used as a baseline measurement, titer is measured using Protein-A affinity chromatography (HPLC). Early in the development of a mAb, a large number of harvest cell culture (HCC) samples must be screened for IgG titer. Affinity chromatography employing a Protein A ligand is often used to determine the mAb concentration as well as to purify it for downstream aggregate and charge variant analysis.

• Doubling rate

  The growth, or doubling rate of the cell line used to produce your mAb is often used as a second decision point by commercial manufacturers like you. The doubling rate reflects the growth potential of the cell line and it influences the amount of mAb product produced as is reflected also by titer.

• Binding affinity to target (surface plasmon resonance)

  Understanding the binding affinity of your mAb with its intended target is an early indicator of generic binding function. Surface plasmon resonance (SPR) is the gold standard in rapid and high throughput binding analysis which gives detailed kinetic information on the nature as well as strength of binding.

• N-glycan analysis

  Highly variable glycosylation may impact mAb purity and produce variable function. Monoclonal antibodies carry N-glycans on their backbone that can be released using LC-MS to assess glycosylation patterns. Characterization of the N-glycans from a monoclonal antibody is necessary as part of a work package providing full structural details of the molecule under investigation. Understanding these N-glycan pathways is important because N-glycans affect many properties of glycoproteins including their conformation, solubility, antigenicity, and recognition by glycan-binding proteins.

• Intact mass

  Determination of the intact molecular mass is a necessary step in characterization of biologics, from clone selection to verification of the final product. Intact molecular mass analysis is commonly used to demonstrate the diversity of the protein/peptide products and verify the identity of the biologics. With optimization, it can also determine the intact molecular weight of all protein products, including bispecific monoclonal antibodies.

• Potency

  Potency assays are essential in verifying product quality: they prove the drug has achieved its desired effect and strength. Potency assays are needed from early development through commercial release because once a drug is ready for market, a different measurement of potency could be reported. This makes it necessary to have the correct potency assay developed as early as possible to ensure safety and avoid delays in your development plan.

• Binding via surface plasmon resonance (SPR) scan or ELISA

  The extensive variability of a molecule's physicochemical and structural properties creates extensive diversity in the Fc region of the antibody, for example through differences in glycosylation profile in the Fc region. Structural differences can influence the binding activity with Fc receptors expressed on immune cells which in turn influences the antibody’s effector function and can therefore have a great impact on potency if effector mechanisms are important for mechanism of action. The Fc region plays a critical role in the function of therapeutic antibodies as they can bind to a variety of receptors and other protein components within the immune system.