Methods in Molecular Biology (2022) 2436: 145–156

DOI 10.1007/7651_2021_411

© Springer Science+Business Media, LLC 2021

Published online: 22 June 2021

Microcarrier-Supported Culture of Chondrocytes

in Continuously Rocked Disposable Bioreactor

Kamil Wierzchowski and Maciej Pilarek

Abstract

Disposable wave-assisted bioreactors are devices originally designed for scaling-up cultures of extremely

fragile animal cells. In such bioreactors, agitation is achieved by continuous horizontal oscillations of

disposable culture bag-like container fixed in a rocker unit. The continuous rocking movement of the

container induces waves in the two-phase (i.e., gas–liquid) culture system composed of CO2-enriched air

and aqueous culture medium. Such continuously oscillating devices can be utilized for supporting homo-

geneity in systems for in vitro propagation of animal anchorage-dependent, that is, adherent, cells, like CP5

chondrocytes cells. As most of in vitro cultured cells exhibit anchorage-dependency toward solid surface,

the suitable interface can be provided by beads of microcarriers made of polymers and characterized by

large surface-to-volume ratio. This chapter describes a methodology for efficient propagation of CP5

chondrocytes on Cytodex 3 microcarriers performed in ReadyToProcess WAVE 25 disposable bioreactor,

as well as all useful procedures for daily monitoring the growth of CP5 chondrocytes.

Key words Anchorage-dependent cells, Bioprocess intensification, Chondrocytes, Disposable biore-

actor, Microcarrier beads, Wave-assisted agitation

1

Introduction

In the case of scaling-up bioprocesses focused on anchorage-

dependent (i.e., adherent) animal cells, the static culture systems

must be substituted by gentle-agitated systems to ensure sufficient

level of aeration in high cell density cultures. Instead of commonly

applied rotating stirrers applied in classical bioreactor systems, con-

tinuously oscillating devices can be utilized for supporting homo-

geneity in disposable (i.e., single-use) bioreactor systems originally

designed for in vitro maintaining of extremely fragile animal cells

[1]. Without a doubt, the disposable bioreactors support notable

reduction of microbial contamination, as well as reduce the devel-

oping cost of the plant constructions, in contrast to typical bior-

eactors equipped with large-volume vessels made of glass or

stainless steel [2, 3]. In the case of single-use bioreactors the agita-

tion is achieved mainly by continuous horizontal oscillations of

disposable culture bag fixed in a rocker unit [4, 5]. The continuous

rocking of the rocker generates, as well as escalates, waves in the

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