Macroporous gelatin microcarriers prepared by calcium carbonate inclusion method were used to cultivate Vero and HepG2 cells. Vero cells attached to these microcarriers at a slightly slower rate and subsequent growth extent was also slightly low compared to their attachment to controlled-charge Cytodex 1 microcarriers.
HepG2 cells, in contrast to Vero cells, do not attach to these microcarriers at a satisfactory rate. Consequently, final cell density was relatively very low and many bared microcarriers were appeared at final stage of the culture. That is poor attachment of some cell type has been impeding their growth on these microcarriers.
Modification of the beads by increasing the charge density through incorporation of (diethylamino) ethylchloride - hydrochloride (DEAE:Cl-HCl) or lysine, drastically improved their attachment and subsequent growth. When microcarriers were modified by addition of 0-2% lysine, positive charge density, negative charge density, and anion exchange capacity were increased from 0.25 meq/g-MC to 0.89 meq/g-MC, from 0.60 meq/g-MC, to 1.0 meq/g-MC, and from 0.38 meq/g-MC to 0.93 meq/g-MC respectively. In case of modification of microcarriers with DEAE:CI-HCl, positive charge density and anion exchange capacity were increased from 0.25 meq/g-MC to 0.60 meq/g-MC and from 0.38 meq/g-MC to 1.7 meq/g-MC, respectivity. But negative charge density was decreased from 0.60 meq/g-MC to 0.40 meq/g-MC.
An increase in charge density of the microcarriers to improve cell attachment may facilitate the cultivation of the cells on macroporous gelatin microcarriers. In case of HepG2 cells, initial attachment rates and attachment yield were markedly increased by increasing charge density. Subsequently, final cell density at confluent state reached to almost $10^7$ cells/ml which was increased by 2-3 times compared with that obtained with normal macroporous gelatin icrocarriers.
The adverse effect of serum proteins on Vero cell attachment was observed. The shielding effect of serum proteins around microcarriers might attenuate the electrostatic force between microcarriers and cells and provent cells from approaching to the microcarriers.
