Tailored Purification Tools for Different Antibody Modalities

Antibody structure Since the 80’s over 100 mAbs have been approved for therapeutic use. Currently, all approved therapeutic antibodies are based on the IgG antibody structure, a ~150 kD protein structure comprised of two identical light chains and heavy chains. Together these chains form a flexible Y-shaped structure. Therapeutic antibody modalities Purification Solutions for Novel Antibody Modalities Therapeutic monoclonal antibodies (mAbs) are customized biopharmaceuticals used for the diagnosis and treatment of various diseases. Traditionally, protein A resins have been considered the workhorse for the purification of mAbs. However, the growing diversity of therapeutic antibody modalities in the drug pipeline has revealed a need for new purification tools. This infographic explores affinity resins specifically designed for the purification of different types of antibody modalities. Learn more about Thermo Scientific™ MabCaptureC and CaptureSelect antibody affinity resins Variable domains (antigen binding) Light chain Fab region Heavy chain IgG antibody Fc region Constant domains Absent or altered Protein A binding site MabCaptureC FcXP FcXP CH1-XL KappaXP LambdaXP MabCaptureC MabCaptureC FcXP FcXP MabCaptureC CH1-XL KappaXP LambdaXP pH sensitive molecules (e.g., bi-specific Ab, Fc-fusion proteins) Formation of dimers and/or aggregation (e.g., bi-specific Ab) Elevated levels of high and low molecular weight species Overexpression of light chains and light chain dimers Antibody-subdomain specific purification Affinity chromatography resins specifically developed to bind antibody-subdomain regions are the solution to these challenges. CaptureSelectTM technology is based on single domain (VHH) antibody fragments immobilized on a chromatograhy matrix. CaptureSelect antibody affinity resins enable increased purity and yield in a single step and are optimized to operate at higher pH compared to Protein A resins. Antibodies can be engineered to create novel therapeutic variants. mAbs are homogenous preparations of antibodies, making them identical in protein sequence, antigen-binding recognition, affinity and downstream functional effects. Find the purification solution that best fits your needs Purification of antibody therapeutics Usually, patients require several grams of a mAb as a clinical dose. Thus, therapeutic mAbs need to be produced at large scale. Over time, production processes, upstream as well as downstream, have been optimized to deliver high yields and to maximize efficiency. Traditionally, Protein A affinity chromatography has been the method of choice for the purification of mAbs. pH • Affinity for the CH1 domain • Purification of all mAbs and Fabs irrespective of light chains • Does not bind free light chains (often product-related impurities) • Affinity for CL-Kappa or CL-Lambda domains • Purification of Fab fragments and bi-specific Abs • High dynamic binding capacity • Mild elution • Affinity for the CH3 domain • Purification of mAbs with altered Protein A binding site and Fc-fusion proteins • Mild elution CaptureSelect™ CH1-XL CaptureSelect™ KappaXP CaptureSelect™ LambdaXP CaptureSelect™ FcXP MabCaptureC Standard antibody Bi-specific antibodies Antibody drug conjugates Fc-fusion proteins Fab fragments MabCaptureC CaptureSelect KappaXP CaptureSelect LambdaXP CaptureSelect FcXP VH VL CL CH1 CH2 CH3 VH VL CL CH1 CH2 CH3 *In order of recommendation Resins for each antibody modality CaptureSelect CH1-XL Standard monoclonal antibody This was the first modality to be used as a therapeutic. It has the basic IgG structure. It binds to a unique epitope of an antigen to activate or inhibit biological processes that are implicated in a specific disease. Antibody Drug Conjugate (ADC) mAb chemically linked to a drug or toxin. ADCs are used to deliver the drug to a specific target protein on a cell. This therapeutic modality is often used to attack cancer cells. Bi-specific antibody Engineered mAbs that can simultaneously bind to two different antigens or two epitopes on the same antigen. The dual functionality opens up a wide range of applications used in a variety of disease pathways. Fc-fusion proteins Fc domain of a mAb is fused to a functional protein. Fc-fusion proteins are often created to prolong half-life of the therapeutic protein. Fc-fusion proteins do not have antigen-binding properties like mAbs have. F(ab) and F(ab’)2 fragments Consists of the variable domain and the first constant domain of each heavy and light chain. These small molecules can enhance pharmacokinetic properties and increase efficiency of tissue or tumor penetration. How do Protein A resins work? Native Protein A is a 55-kDa bacterial cell wall protein that naturally binds to IgG molecules. Because of these binding properties, Protein A can be used as a ligand on a chromatography support and used for the purification of mAbs. The MabCaptureC affinity matrix is a high-performance Protein A resin specifically designed to improve the purification efficiency of mAbs with intact Protein A binding sites. Purification challenges of novel antibody modalities The development of novel antibody modalities introduced some challenges in the purification process making Protein A affinity resins not always effective. VL CL CH1 CH2 CH3 MabCaptureC MabCaptureC CL CH1 CH3 CH3 CH3 CH3 CH1 CH1 CL CL