Comparison of different ligand densities in immunoaffinity chromatography of the plantibody HB-01 coupled to Sepharose CL-4B to purify the rHBsAg

Valdés, Rodolfo; Medina, Yenisley; Ferro, William; Reyes, Biunayki; Geada, Déborah; Montero, José; Alvarez, Tatiana; Leyva, Alejandro; Gómez, Leonardo D.; Padilla, Sigifredo; Pacín, Leonardo; Figueroa, Alejandro; Tamayo, Andrés; Milá, Lorely; Aldama, Yurisleydi; Moya, Galina; Reonde, Jorge; Abrahantes, María del Carmen

doi:10.1016/j.jchromb.2006.12.018

Cited by 2 publications

(1 citation statement)

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“…Currently, one of the technical bottlenecks limiting commercial-scale production of plant-derived antibodies (Abs) and other bioproducts is the high cost and inefficiency of downstream processing and purification . Purification of monoclonal antibodies (mAbs) and Ab fragments from plants relies on commercial chromatography reagents such as Streamline protein A , protein A magnetic beads , expanded-bed adsorption-immobilized metal affinity chromatography (EBA-IMAC) , , protein G-agarose , or combinations thereof . High levels of plant polysaccharides and phenolic compounds can interfere with purification , , and from our experience poorly clarified plant homogenates can lead to resin fouling, requiring considerable maintenance time (J.C.H., unpublished data).…”

Section: Introductionmentioning

confidence: 99%

Purification of Plant-Derived Antibodies through Direct Immobilization of Affinity Ligands on Cellulose

Hussack

Grohs

Almquist

et al. 2010

J. Agric. Food Chem.

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Plants possess enormous potential as factories for the large scale production of therapeutic reagents such as recombinant proteins and antibodies. A major factor limiting commercial advances of plant-derived pharmaceuticals is the cost and inefficiency of purification. As a model system, we have developed a simple yet robust method for immobilizing affinity capture ligands onto solid supports by interfacing the secreted expression and coupling of a chimeric fusion protein in Pichia pastoris to microcrystalline cellulose in a single step. The fusion protein, which consisted of antibody-binding proteins L and G fused to a cellulose-binding domain (LG-CBD), was tethered directly onto cellulose resins added to P. pastoris cultures and subsequently used for antibody purification. Both the antibody-binding protein L and protein G domains were functional, as demonstrated by the ability of cellulose-immobilized LG-CBD to purify both a scFv antibody fragment from yeast and a human IgG1 monoclonal antibody from transgenic tobacco. Furthermore, combining two P. pastoris strains expressing LG-CBD and scFv with CP-102 cellulose in a single culture allowed for easy recovery of biologically active scFv. Direct immobilization of affinity purification ligands, such as LG-CBD, onto inexpensive support matrices such as cellulose is an effective method for the generation of functional, single-use antibody purification reagents. Straightforward preparation of purification reagents will help make antibody purification from genetically modified crop plants feasible and address one of the major bottlenecks facing commercialization of plant-derived pharmaceuticals.

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