The global market for biologics, such as monoclonal antibodies and antibody-drug conjugates, is growing. However, developing and manufacturing these essential drugs is costly, and the risk of failure can be high. By addressing common cell-line development (CLD) challenges, advanced flow cytometry promises to accelerate the drug discovery process and bring high-quality biologics to patients sooner.
Cell-line development begins with cloning antibody genes into an expression vector, which is then used to transfect a mammalian host cell line. Chinese hamster ovary (CHO) cells are widely used for biologics production as they are relatively easy to culture, provide high yields, and can replicate the post-translational modifications and folding found in human proteins.
Next, stably-transfected single cells are isolated for clonal expansion, allowing for assessment of parameters including antibody titer, viability, and growth. Once high-productivity clones with optimal critical quality attributes (CQAs) and target specificities have been identified, culture conditions and media formulations must be tailored to ensure peak cell growth and protein production.
Following this, one or more of the best-performing clones is characterized for stability, productivity, and key antibody properties. For example, antibody binding kinetics are often measured with label-free techniques, using a system such as the Octet® bio-layer interferometry (BLI) system. Finally, the cells are banked in accordance with good manufacturing practices (GMP), ready for scale-up and commercial manufacture.
An initial challenge for CLD lies in choosing a cell line that can create all the biological features of the antibody therapeutic. Factors to consider include stability, growth characteristics, and media requirements, as well as whether the cell line has a documented history.
Identifying the best-performing clones is another difficult step. While screening large numbers of clones can improve the likelihood of discovering a rare, highly productive cell line, increased sample numbers can extend the timeline for process development. This can be especially apparent when samples must be sent away to an external provider for analysis.
The next hurdle is characterization. Comprehensive bioanalysis is crucial to ensure product safety and efficacy, but often involves repeated sampling, running diverse assays across several instruments, and integrating datasets for analysis. This uses up precious samples and also takes time.
In addition, scaling up production can introduce more problems since early-stage models may not accurately represent conditions at larger scales.
Advanced flow cytometry streamlines the process of identifying and characterizing high-performing clones by assessing phenotype and function at greater speed, depth, and scale.
To put this into context, consider that a traditional CLD workflow might involve immunophenotyping cells in tubes with a standard flow cytometer, evaluating viability using a plate-based assay, and measuring antibody titers by ELISA, before piecing all the different datasets together. This could equate to days or even weeks spent on clone evaluation, and often limits the number of parameters analyzed per sample.
In contrast, advanced flow cytometry systems combine multiple readouts in the same well of a 96- or 384-well plate. Not only does this reduce sample and reagent requirements, allowing opportunities to run more replicates, but it also increases throughput and makes optimization more efficient since researchers can obtain a broader overview of each clone in less time.
Furthermore, by eliminating the need to export data for analysis, advanced flow cytometry systems minimize the risk of error that could lead to unnecessary delays.
The iQue® Advanced Flow Cytometer is a next-generation flow cytometry platform with utility for CLD. With the capacity to analyze up to 15 parameters per clone, using as little as 5 µL of sample (in a 384-well plate), it provides phenotypic and functional information in a rapid, high-throughput manner—just 15 minutes to screen an entire 96-well plate. Used in conjunction with multiplex products including the iQue® Cell Count and Viability Kit and the iQue® Human IgG Titer and Viability Kit, the iQue® platform supports rapid clone evaluation and better clone selection to accelerate CLD and shorten time to market.
To learn more about the iQue® platform and how it can accelerate your CLD workflow, visit www.sartorius.com.