DoE-Assisted Method Development for the Analysis of Monoclonal Antibodies Charge Variants by Cation-Exchange Chromatography

[#1]

Cation-exchange chromatography (CEX) is the industry gold standard for the analysis of biopharmaceuticals charge variants such as monoclonal antibodies (mAbs). However, the development of CEX methods in a time and resource-efficient manner constitutes a bottleneck in protein characterisation. CEX separations are complex and governed by multiple factors such as column chemistry, gradient type, flow rate, mobile phase composition and pH. Several scientific publications have proven the successful application of design-of-experiment (DoE) in chromatography method development. Nevertheless, performing DoEs with a large number of factors may be challenging, time-consuming and expensive. This work illustrates the use of a split-DoE approach to aid the development of a CEX method for the analysis of the charge variants profile of a mAb candidate. Analytical method development was intended to provide a high-throughput (HT) CEX method to support charge variants analysis with minimal sample and time requirements. The split-DoE approach is based on fundamental knowledge of the CEX separation mechanism and aims to reduce the number of experimental runs whilst exploring a wide experimental space. Regression modelling was used to study the effect of both individual process parameters and their interactions on the separation efficiency to ultimately identify the optimal method conditions. This study provides an efficient workflow for leveraging the development of CEX methods.

[#2]

Hi @jlmbs,

Thanks a lot for sharing your interesting use case !
I may be mistaking, but I think the design shown under Central Composite Design (pages 5 and looks more like a Box Benhken design with a center point.
A CCD would look like this : https://www.researchgate.net/profile/Harald-Hetzner/publication/275971176/figure/fig1/AS:61429887535...

In your study, did you use a CCD (through augmentation of the Main Effects Screening design) or a "new" Box Benhken design ?
Thanks a lot for your answer, 

[#3]

Hi Victor,

You are right, the cube represents a box Behnken design (BBD) whilst the experiment for the first molecule reported in the slides was done on a central composite design (CCD). Thanks for spotting it and apologies for the typo! BBD was used for optimising CEX methods for other molecules with narrower operating zone for the same experimental factors. The split-doe approach for CEX method development using either a BBD or CCD in the second step was used for the molecules of which CEX traces are reported in slide 9. Top used CCD and bottom used BBD as second DoE. I hope this answers your question and apologies for the confusing slide deck.