Manufacture Of Collagen Carriers And Its Application In CHO Cell Culture

Animal cell culture, by which many bio-products and bio-pharmacies aremanufactured such as monoclonal antibodies, vaccines, recombinant proteins and soon, plays more and more important role in biotechnology and bioengineering. For itsvital character and great profits, a great deal of currents and human resources areabsorbed and great rewards for their efforts have been made. Study on bioreactors andcarriers for animal cell culture has drawn great attentions for their vital status in thisfield. For its high bio-compatibility and stability to chemical and physical factors,collagen is widely applied in industries such as medicine and food.In this paper, the methods to manufacture new carriers with collagen areinvestigated. The main procedure is to denature natural casing by chemical reagents,just like chromium, formaldehyde, glutaraldehyde, chromium-glutaraldehydecomplex and aluminum-tannin complex etc, to improve its heat resistance, stability tochemical reagent, membrane flux and bio-compatibility. Then, the comparison andevaluation of performance and characteristic for culturing CHO cells by the newcarrier made by above modified natural casing, A-DISK carrier imported from U.S.A.and C-DISK carrier manufactured by ourselves have been made. Some estimation ontheir bio-compatibility is given for the new carrier made by above modified naturalcasing based on the experiment data of cell anchorage, glucose metabolic rate andEPO secreted concentration. Furthermore, the principle of cell culture is discussedaccording to metabolic rate of glucose and EPO concentration in medium.The following conclusions are drawn from all of the experiments:1. Natural casing membrane denatured by glutaraldehyde keeps its characters afterheating, and has better surface properties. Moreover, it could resist stream sterilizationand keep its performance stable, which makes casing membrane denatured byglutaraldehyde fit for cell carriers.2. Natural casing membrane denatured by glutaraldehyde show the highest flux toBSA and lysozyme, its flux to BSA is 2.47 mg·cm-2·h-1, its flux to lysozyme is 7.26mg·cm-2·h-1 , better than those nature and other denatured casing membranes. i3. The anchorage experiments of CHO cells on carriers show that natural casing membranedenatured by glutaraldehyde can be firmly stuck by CHO cells, also longer culture time doesno infection on cells' anchorage.4. The highest metabolic rate of glucose in the culture procession on casing membranedenatured by glutaraldehyde is 14.2 mmol?ml-1?h-1, which is improved by 16 %higher than that of C-DISK and 13 % than that of A-DISK, EPO concentration is 282U/ml on casing membrane denatured by glutaraldehyde, which is improved 30 %higher than that of C-DISK and 18 % than that of A-DISK.5. The change of metabolic rates of glucose in cell culture reveals different metabolic level andprotein expression level, but they are not in synchronization. In our experiments, there is about 1-2days delay period between the change of glucose metabolic rate and EPO concentration, thisvacancy mean that we could inspect and detect the culture station.6. The cost of the carrier made by casing membrane denatured by glutaraldehyde isabout 2.0 RMB/g, which is about 5 % of A-DISK; the cost of C-DISK is 0.2 RMB/g,only 0.5 % of A-DISK.Although both denatured casing carrier and C-DISK carrierare much cheaper than imported A-DISK carriers, denatured casing carrier can still beregarded as the best carrier for their advantage in cell growth, protein expression.