Cell clarification represents a major challenge for the intensification through very high cell density in the production of biopharmaceuticals such as monoclonal antibodies (mAbs). The present report proposes a solution to this challenge in a streamlined process where cell clarification and mAb capture are performed in a single step using magnetic beads coupled with protein A. Capture of mAb from non-clarified CHO cell suspension showed promising results; however, it has not been demonstrated that it can handle the challenge of very high cell density as observed in intensified fed-batch cultures. The performances of magnetic bead-based mAb capture on non-clarified cell suspension from intensified fed-batch culture were studied. Capture from a culture at density larger than 100 × 106 cells/ml provided an adsorption efficiency of 99% and an overall yield of 93% with a logarithmic host cell protein (HCP) clearance of ≈2–3 and a resulting HCP concentration ≤≈5 ppm. These results show that direct capture from very high cell density cell suspension is possible without prior processing. This technology, which brings significant benefits in terms of operational cost reduction and performance improvements such as low HCP, can be a powerful tool alleviating the challenge of process intensification. 

High capacity magnetic protein A agarose beads, LOABeads PrtA, were used in the development

of a new process for affinity purification of monoclonal antibodies (mAbs) from non-clarified

CHO cell broth using a pilot-scale magnetic separator. The LOABeads had a maximum binding

capacity of 65 mg/mL and an adsorption capacity of 25–42 mg IgG/mL bead in suspension for an

IgG concentration of 1 to 8 g/L. Pilot-scale separation was initially tested in a mAb capture step

from 26 L clarified harvest. Small-scale experiments showed that similar mAb adsorptions were

obtained in cell broth containing 40 Å~ 106 cells/mL as in clarified supernatant. Two pilot-scale

purification runs were then performed on non-clarified cell broth from fed-batch runs of 16 L,

where a rapid mAb adsorption ≥96.6% was observed after 1 h. This process using 1 L of magnetic beads had an overall mAb yield of 86% and 16 times concentration factor. After this single protein

A capture step, the mAb purity was similar to the one obtained by column chromatography, while

the host cell protein content was very low, <10 ppm. Our results showed that this magnetic bead

mAb purification process, using a dedicated pilot-scale separation device, was a highly efficient

single step, which directly connected the culture to the downstream process without cell clarification.

Purification of mAb directly from non-clarified cell broth without cell separation can provide

significant savings in terms of resources, operation time, and equipment, compared to legacy procedure of cell separation followed by column chromatography step.

CD133+ cells are potential myogenic progenitors for skeletal muscle regeneration to treat muscular dystrophies. The proliferation of human CD133+ stem cells was studied for 14 days in 3D biomimetic electrospun poly-L-lactic acid (PLLA) nano-fiber scaffolds. Additionally, the myogenic differentiation of the cells was studied during the last 7 days of the culture period. The cells were homogeneously distributed in the 3D scaffolds while colony formation and myotube formation occurred in 2D. After a lag phase due to lower initial cell attachment and an adaptation period, the cell growth rate in 3D was comparable to 2D after 7 and 14 days of culture. The expression of the stem cell (SC) marker PAX7 was 1.5-fold higher in 3D than 2D while the differentiation markers MyoG, Desmin and MyoD were only slightly changed (or remain unchanged) in 3D but strongly increased in 2D (12.6, 3.9, and 7.9-fold), and the myotube formation observed in 2D was absent in 3D. The marker expression during proliferation and differentiation, together with the absence of myotubes in 3D, indicates a better maintenance of stemness in 3D PLLA and stronger tendency for spontaneous differentiation in 2D culture. This makes 3D PLLA a promising biomaterial for the expansion of functional CD133+ cells.