Methods in Molecular Biology (2022) 2436: 1–15

DOI 10.1007/7651_2021_441

© Springer Science+Business Media, LLC 2021

Published online: 06 October 2021

Benchtop Bioreactors in Mammalian Cell Culture:

Overview and Guidelines

Ange´lique Schmid, Emanuel Kreidl, Martin Bertschinger,

and Patrick Vetsch

Abstract

Bioreactors are manufactured apparatuses that allow the generation of a specific environment for the highly

controlled cultivation of living cells. Originally used for microbial production systems, they have found

widespread applications in fields as diverse as vaccine production, plant cell cultivation, and the growth of

human brain organoids and exist in equally diverse designs (Chu and Robinson, Curr Opin Biotechnol 12

(2):180–187, 2001; Qian et al., Nat Protoc 13:565–580, 2018). Manufacturing of biologics is currently

mostly performed using a stirred tank bioreactor and CHO host cells and represents the most “classical”

bioreactor production process. In this chapter, we will therefore use the cultivation of suspension Chinese

hamster ovary (CHO) cells for recombinant protein production in a stirred tank bioreactor as an example.

However, general guidelines provided in this chapter are transferable to different bioreactor types and host

cells (Li et al., MAbs 2(5):466–479, 2010).

The preparation and operation of a bioreactor (also referred to as upstream process in a biotechnologi-

cal/industrial setting) is comprised of three main steps: expansion (generation of biomass), production

(batch, fed-batch, or continuous process), and harvest. The expansion of cells can last from few days to

weeks depending on the number of cells at the start, the cellular doubling time, and the required biomass to

inoculate the production bioreactor. The production phase lasts a few weeks and is a highly sensitive phase

as the concentration of different chemicals and physical parameters need to be tightly controlled. Finally,

the harvest will allow the separation of the product of interest from large particles and then the desired

material (cell culture supernatant or cells) is transferred to the downstream process.

The raw materials used during the upstream phase (all three steps) need to be aligned with the final

purpose of the manufactured product, as the presence of residual impurities may have an impact on

suitability of the final product for a desired purpose.

Key words Bioreactor, Mammalian cell culture, Feeding strategy, Fed-batch, Recombinant protein

production

1

Introduction

For the production of complex therapeutic proteins like recombi-

nant antibodies, mammalian cell lines have been the workhorse of

the industry for three last decades [1, 2]. Chinese hamster ovary

(CHO) cells in particular have found widespread application as they

have a history of safe use, they exhibit a robust growth capacity with

reasonable doubling times, they can be readily modified to express

1