Affinity Perfusion Chromatography

Applications of affinity chromatography for quantitative analysis (
1
–
10
), purification at laboratory scale (
11
–
16
), and for large-scale manufacture of recombinant
DNA
technology derived therapeutics (
17
–
23
), have been continually expanding since the first application introduced by Cuatrecasas in 1968 (
24
). Although there have been some examples of the use of rigid high-performance liquid chromatography (HPLC)-based supports for affinity chromatography, the majority of applications are found on agarose-based particles. The historic utility of agarose as an affinity support stems from attributes such as a well-developed base of activated chemistries, relatively large pore volume for immobilization of proeteinaceous ligands, chemical inertness, and charge neutrality (
25
–
29
). However, the particle porosity is defined by the degree of swelling and therefore, the holdup of solvent acts as a stagnant pool inside the particle. This solvent pool leads to slow mass transport to interior binding sites, necessitating relatively low operating flow rates and long cycle times (
30
–
33
). For many years, these attractive features, overcame the inherently slow speed of operation dictated by these soft gel columns.