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BMC Proceedings

Open Access

Use of microcarriers in Mobius® CellReady bioreactors to support growth of adherent cells

  • Michael McGlothlen1Email author,
  • Donghui Jing1,
  • Christopher Martin1,
  • Michael Phillips1 and
  • Robert Shaw1
BMC Proceedings20137(Suppl 6):P95

https://doi.org/10.1186/1753-6561-7-S6-P95

Published: 4 December 2013

Abstract

The Mobius® CellReady bioreactor product platform incorporates novel disposable technologies that provide optimal performance for suspension mammalian cell culture. Here we show the utility of EMD Millipore's 3L and 50L CellReady single use bioreactors for the cultivation of adherent mammalian cells on microcarriers. Cytodex3® and Solohill® collagen microcarriers were first tested in a mixing study to assess feasibility. We evaluated the normalized mixing speed required in the 3L and 50L to achieve a suspension of the microcarriers and enable growth of the cells.

Mixing

Manufacturer specifications show Cytodex3® and Solohill® microcarriers to be similar in density and size. Working with this assumption, mixing studies where performed using the Cytodex3® microcarriers in 3L Mobius® CellReady and Solohill® Collagen coated in 50L single use bioreactor to determine the slowest agitation speed or the just suspended mixing power inputs (P/V)js, required to fully suspend the microcarriers so that the beads are equally distributed in the bioreactor.

Microcarrier distribution was assessed by sampling the bioreactor at varying depths. Then the dry weight of the microcarrier was used to determine the% relative sample weight to the target weight.

Mixing Results

Data show the (P/V)js to be ~0.6W/m3 in both the 3L and 50L single use bioreactors

100% distribution corresponds to the theoretical concentration of microcarriers, which is 3g/L Cytodex3® in 3L bioreactor and 15g/L Solohill® Collagen microcarriers in 50L bioreactor

Cell Growth

Initial cell culture runs were performed with MDCK and Human Mesenchymal Stem Cells (hMSCs) to evaluate the bioreactor agitation to support cell growth in the 3L Mobius® CellReady single use bioreactor. The conditions that showed the best performance could then scaled to the 50L Mobius® bioreactor.
  1. 1.

    Cultured MDCK cells on Cytodex3® microcarriers grew to a peak cell density of ~1e6cells/mL using a power input of 0.6W/m3 with a 2L working volume after 3 days.

     
  2. 2.

    Cultured hMSCs on Solohill® microcarriers grew to a maximum total cell number of 6e6 cells using power input of 0.6-0.8W/m3 with a 2.4L working volume after 12 days.

     

Conclusions

  1. 1.

    Data from the mixing experiments demonstrate the just suspended mixing power input was determined to be ~0.6W/m3.

     
  2. 2.

    Cell growth experiments with hMSCs demonstrate comparable cell growth in the 3L and 50L Mobius® CellReady bioreactor with total number of hMSCs reaching 4e8 and 9e9 cells after 12 days at a agitation power input of 0.6-0.8W/m3

     
  3. 3.

    Initial cell growth experiments with adherent MDCK cells demonstrate an agitation power/volume input of 0.6W/m3 may provide the best performance for cell growth with peak cell densities ~1.0e6 cells/mL after 3 days

     
  4. 4.

    Comparable MDCK cell growth is observed:

     

Mobius® CellReady Bioreactor 3L

Mobius® CellReady Bioreactor 50L

Rocking Bioreactor 20L
Figure 1

Illustrates the attachment of MDCK and hMSCs to Cytodex 3 ® and Solohill ® microcarriers

Figure 2

compares the viable cell density of MDCK cells at increasing power/volume impeller inputs and different bioreactors

Figure 3

shows the viable cell density of hMSCs in the 3L and 50L Mobius ® CellReady Bioreactor

Table 1

Physical Characteristics of Microcarriers

Microcarrier

Cytodex 3 ®

Solohill® Collagen Coated

Density (g/ml)

1.04

1.03

Hydrated Size (µm)

141-211

125-212

Concentration (g/ml)

3

15

Table 2

MDCK/ Cytodex 3 ® Microcarriers Process Table

Variable

Value

Cells

MDCK

hMSCs

Inoculation Density

4e5 cells/mL

5e3 cells/mL

Substrate

Cytodex 3 ®

Solohill®

Growth Media

DMEM w/ 4.5g/L Glucose, 2% FBS, 1% NEAA and 2mM L-Glutamine

DMEM low glucose, 10% FBS, 8ng/ml bFGF, 2mM Glutamine, 1X Pen/Strep

pH

7

NA

DO (% Saturation)

45

NA

Feed 1

Day 1: 100% Growth Media

Day 6: 1000ml low glucose fresh medium

Feed 2

Day 3: Drain 50% of the working volume and reefed with equal volume of Growth Media

Day 9: 400ml high glucose fresh medium

Batch Duration

7 days

12 days

Authors’ Affiliations

(1)
EMD Millipore Corporation

Copyright

© McGlothlen et al; licensee BioMed Central Ltd. 2013

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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