Two-Dimensional Liquid Chromatography Applications in Biopharmaceutical Analysis

Biopharmaceutical analysis typically involves the use of numerous, orthogonal, high-resolution analytical tools. As such, analytical characterization of biotherapeutic products is a resource-intensive activity and is in focus currently as biopharmaceutical manufacturers look to make development of these products cost-effective. In this context, two-dimensional liquid chromatography (2D-LC) has emerged as an efficient multidimensional approach for biopharmaceutical analysis. As proteins are inherently complex molecules, they offer significant analytical challenges owing to structural variations, post-translational modifications (PTMs), or the tendency to aggregate. Traditional one-dimensional liquid chromatography (1D-LC) methods have long been used to efficiently separate these biomolecules. However, over the past few years, 2D-LC has demonstrated superior separation capacity and resolution, garnering a spectrum of applications for entities such as monoclonal antibodies (mAbs), antibody-drug conjugates (ADCs), and other therapeutic proteins. Development of 2D-LC methods offers plentiful opportunities for innovation in instrumentation, automation, and data analysis. This article reviews literature related to 2D-LC applications in the biopharmaceutical field over the past six years (2018-2024).