Serum-free suspension culturing of human cells: adaptation, growth, and cryopreservation

Author(s)

Biaggio, Rafael Tage ^[1] ; Abreu-Neto, Mario Soares ^[2] ; Covas, Dimas Tadeu ^{[2, 3]} ; Swiech, Kamilla ^[1]

Affiliation

^[1] Univ Sao Paulo, Sch Pharmaceut Sci Ribeirao Preto, BR-14040903 Sao Paulo - Brazil

^[2] Hemotherapy Ctr Ribeirao Preto, BR-14051140 Sao Paulo - Brazil

^[3] Univ Sao Paulo, Ribeirao Preto Med Sch, BR-14049900 Sao Paulo - Brazil

• Document type: Journal article
• Source: Bioprocess and Biosystems Engineering; v. 38, n. 8, p. 1495-1507, AUG 2015.
• Web of Science Citations: 6

Abstract

Human cell lines have attracted great interest because they are capable of producing glycosylated proteins that are more similar to native human proteins, thereby reducing the potential for immune responses. However, these cells have not been extensively characterized and cultured under serum-free suspension conditions. In this work, we describe the adaptation, growth, and cryopreservation of the human cell lines SK-Hep-1, HepG2, and HKB-11 under serum-free suspension conditions. The results showed that both HKB-11 and SK-Hep-1 adapted to serum-free suspension cultures in FreeStyle and SFM II, respectively. Kinetic characterization showed that the HKB-11 and SK-Hep-1 cells reached cell densities as high as 8.6 x 10(6) and 1.9 x 10(6) cells/mL, respectively. The maximum specific growth rates (mu (max)) were similar for both cells (0.0159/h for HKB-11 and 0.0186/h for SK-Hep-1). The growth limitation of adapted cells does not appear to be associated with glucose or glutamine depletion, nor with the formation of lactate in inhibitory concentrations. However, in both cases, ammonia production reached concentrations that are considered inhibitory to mammalian cells (2-5 mM). The adapted cells were also successfully cryopreserved under serum-free formulations. The SK-HEP-1 and HKB-11 cells that were adapted to serum-free suspension conditions might be suitable for use in the manufacturing of recombinant proteins, thereby eliminating the potential for the introduction of adventitious process contamination and greatly simplifying downstream protein purification. (AU)

• FAPESP's process: 12/04629-8 - Establishment of a production platform for recombinant therapeutic proteins in human cells
• Grantee: Kamilla Swiech Antonietto
• Support Opportunities: Research Grants - Young Investigators Grants
• FAPESP's process: 12/02109-7 - SERUM-FREE SUSPENSION ADAPTATION OF HUMAN CELL LINES
• Grantee: Rafael Tagé Biaggio
• Support Opportunities: Scholarships in Brazil - Master