Engineering characterisation of single-use bioreactor technology for mammalian cell culture applications

Abstract

Background The commercial success of mammalian cell-derived recombinant proteins has fostered an increase in demand for novel single-use bioreactor (SUB) systems, that facilitate greater productivity, increased flexibility and reduced costs. Whilst maintaining auspicious mixing parameters, these systems exhibit fluid flow regimes unlike those encountered in traditional glass/stainless steel bioreactors. With such disparate mixing environments between SUBs currently on the market, the traditional scale-up procedures applied to stirred tank reactors (STRs) are not adequate. The aim of this work is to conduct a fundamental investigation into the hydrodynamics of single-use bioreactors at laboratory scale to understand its impact upon the growth, metabolic activity and protein productivity of an antibody-producing mammalian cell culture. Materials and methods This work presents a study characterising the macromixing, fluid flow pattern, turbulent kinetic energy (TKE), energy dissipation rates (EDRs), and shear stresses within these bioreactor systems carried out using 2dimensional Particle Image Velocimetry (PIV). PIV enables acquisition of whole-field flow characteristics through instantaneous velocity measurements. The SUBs employed in the PIV measurements include the 3L CellReady (Merck Millipore), PBS Biotech’ sP BS 3 bioreactor and the Sartorius 2L BIOSTAT Cultibag RM. The CellReady is a stirred tank bioreactor (3 litre volume), housing a 3-bladed upward-pumping marine scoping impeller. The PIV study was conducted using the actual vessel which has an internal diameter (DT )o f 137mm and height (HT) of 249mm. The marine scoping impeller (DI) is 76.2mm in diameter and is located near the bottom with a clearance of 30mm from the base. Measurements were obtained at varying impeller rates from 80 to 350rpm (corresponding to Re =8 699 to 38057). The PBS 3 is a pneumatically driven bioreactor (3 litre volume) whose mixing is induced through the buoyancy of bubbles. PIV measurements were again obtained utilising the actual PBS 3 vessel in the central vertical plane of the bioreactor at wheel speeds of 20, 27, 33 and 38rpm. The Sartorius Cultibag RM is a rocked bag bioreactor with a 2 litre total volume. A custom-made Sartorius Cultibag mimic and rocking platform was manufactured to enable the required optical access for PIV investigations. Measurements were taken at a rocking speed of 25rpm, in the vertical plane 8.5cm from the outer edge of the bioreactor. Fluid working volume (wv) was varied at 30, 40, 50 and 60% wv. A biological study into the impact of these fluid dynamic characteristics on mammalian cell culture performance and behaviour is presented. CellReady and Cultibag cell cultures were conducted using the GSCHO cell-line (Lonza) producing an IgG4 (B72.3) antibody. The impeller speed and working volume are used to vary the hydrodynamic environment within the CellReady, whilst the rocker speed is the varied parameter in the Cultibag RM. Results and discussion The upward-pumping 3-bladed impeller within the CellReady engenders compartmentalisation of the fluid flow. This in turn contributes to the wide range of turbulence levels conveyed between the lower quarter and upper three quarters of the fluid. The maximum fluid velocity of 0.25Utip is achieved in the impeller discharge stream