Influence of different spacer arms on Mimetic Ligand™ A2P and B14 membranes for human IgG purification

Microporous membranes are an attractive alternative to circumvent the typical drawbacks associated to bead-based chromatography. In particular, the present work intends to evaluate different affinity membranes for antibody capture, to be used as an alternative to Protein A resins. To this aim, two Mimetic LigandsTM A2P and B14, were coupled onto different epoxide and azide group activated membrane supports using different spacer arms and immobilization chemistries. The spacer chemistries investigated were 1,2-diaminoethane (2LP), 3,6-dioxa-1,8-octanedithiol (DES) and [1,2,3] triazole [TRZ]. These new mimetic membrane materials were investigated by static and by dynamic binding capacity studies, using pure polyclonal human immunoglobulin G (h-IgG) solutions as well as a real cell culture supernatant containing monoclonal h-IgG1. The best results were obtained by combining the new B14 ligand with a TRZ-spacer and an improved Epoxy 2 membrane support material. The new B14-TRZ-Epoxy 2 membrane adsorbent provided binding capacities of approximately 3.1 mg/mL, besides i) a good selectivity towards IgG, ii), high IgG recoveries of above 90%, iii) a high Pluronic-F68 tolerance and iv) no B14-ligand leakage under harsh cleaning-in-place conditions (0.6 M sodium hydroxide). Furthermore, foreseeable improvements in binding capacity will promote the implementation of membrane adsorbers in antibody manufacturing.