Development of halal gelatin immobilized polystyre microcarriers and its performance in vero cell culture /
Microcarriers (MCs) serve as the substrates in bioreactor for attractive cell proliferation of anchorage-dependent cells. Surface engineering of MCs is a critical effort for tuning the interactions of cells to the engineered matrix. The present work aims to develop biocompatible polymeric MCs to obt...
Saved in:
| Main Author: | |
|---|---|
| Format: | Thesis |
| Language: | English |
| Published: |
Kuala Lumpur:
Kulliyyah of Engineering, International Islamic University Malaysia,
2012
|
| Subjects: | |
| Online Access: | http://studentrepo.iium.edu.my/handle/123456789/4628 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| Summary: | Microcarriers (MCs) serve as the substrates in bioreactor for attractive cell proliferation of anchorage-dependent cells. Surface engineering of MCs is a critical effort for tuning the interactions of cells to the engineered matrix. The present work aims to develop biocompatible polymeric MCs to obtain a higher Vero cell density in suspension culture. MCs were developed from microsize hydrophobic polystyrene (PS) beads ranging from 100-150 μm which were surface modified by ozone aeration (O3), a combination of ozone aeration and UV irradiation (UVO3) and a combination of ozone aeration and UV irradiation in the presence of water (UVO3W). An increase in surface hydrophilicity and functionality of the PS beads upon surface oxidation by the introduction of carbonyl (CO), hydroxyl (OH) and carboxyl group (COOH) was confirmed by FTIR and XPS spectra. Highest COOH functional group and better surface wettability was introduced by UVO3 treatments. Following the surface treatment, covalent immobilization of halal bovine gelatin on the treated PS by a combination of 1 ethyl 3- (3-dimethylaminopropyl carbodiimide) (EDAC) and N-hydroxysuccinimide (NHS) obtained the highest and stable amount of immobilized gelatin with the presence of amide bond (CONH and NH) and an improved thermal properties compared to and N, N' carbonyldiimidazole (CDI) or EDAC alone. Results found that moderate contact angle value ranging from 67⁰-68⁰ and COOH density of 0.17224 x 10-6 mmol/mg at an increased surface roughness provides a preferable surface for gelatin coupling. Optimization study found that 45 min treatment time at 1.5 L/min flowrate and 0.85 mW/cm2 is the most significant conditions to modify PS with respect to covalent immobilization of gelatin. The density of beads of 1.02-1.03 g/cm3 was slightly higher than the culture media which enables suspension of MCs in the spinner vessel culture. Immobilizations of halal gelatins (bovine, fish and its cationized forms) on treated PS were prepared and were evaluated for their ability to support Vero cells expansion. Interestingly, it was observed that the rejection of Vero cells on positively charge substrate demonstrated by poor growth of Vero cells on cationized bovine gelatin (CBG) and cationized fish gelatin (CFG). Study found that zeta potential in the range of -20 to -24mV promoted better cell attachment and growth. Covalently immobilized bovine gelatin on polystyrene (BGPS) is an effective MCs having excellent surface for cell attachment and proliferation, easy sampling and easy cell recovery, good microscopic and suspension ability with highest maximum cell number of 8.0 x 107 cells quantified at 4g/L MCs concentration, 60 rpm agitation speed and 0.8 x 105 cell/mL inoculum density. Three levels factorial design under the Response Surface Method (RSM) suggested that each independent parameter is highly interrelated and will not perform at best in the absence of the other. A comparative study with two commercial polystyrene MCs from Solo Hill Engineering revealed that BGPS is better at the optimum culture condition. The experimental outcomes demonstrated that halal MCs have been successfully developed with UVO3 apparatus which is a convenient and reliable method for modification of the PS microparticles for subsequent covalent immobilization of halal bovine gelatin and that the resulting engineered beads are suitable for MCs applications. |
|---|---|
| Item Description: | Abstract in English and Arabic. "A thesis submitted in fulfilment of the requirement for the degree of Master of Science (Biotechnology Engineering)."--On t.p. |
| Physical Description: | xxiv, 239 leaves : ill. ; 30cm. |
| Bibliography: | Includes bibliographical references (leaves 184-219). |