Wongso, Hanny Angelique and Tunjung Mahatmanto, STP., M.Si., PhD and Yunita Tirtasari Soenaryo, S.Si, M.Si (2024) Optimasi Seeding Density pada Proses Produksi Recombinant Human Erythropoietin. Sarjana thesis, Universitas Brawijaya.
Abstract
Produksi recombinant human erythropoietin (EPO) menggunakan sel Chinese Hamster Ovary (CHO) adherent dapat dioptimasi dengan menentukan seeding density yang optimum. Seeding density sebagai jumlah sel awal pada proses kultur akan mempengaruhi pertumbuhan dan produktivitas sel, ketersediaan nutrisi, proses glikosilasi, hingga karakteristik EPO yang dihasilkan. Saat ini, proses kultur dalam produksi EPO masih menggunakan split ratio dimana jumlah sel yang diinokulasi dapat bervariasi. Hal ini dapat mempengaruhi konsistensi proses dan kualitas produk akhir sehingga penentuan seeding density optimum perlu dilakukan. Studi ini membandingkan penggunaan seeding density dengan perlakuan 3×107 (A), 4×107 (B), 5×107 (C), 6×107 (D), dan 7×107 (E) sel. Sel dikultur hingga tahap produksi menggunakan serum-free media, lalu supernatan dipanen dan dipurifikasi hingga tahap kromatografi IEX2. Kemudian, analisis dan karakterisasi EPO dari semua perlakuan dilakukan. Hasil penelitian menunjukkan bahwa seeding density memiliki pengaruh terhadap produktivitas dan karakteristik EPO. Hasil analisis kadar sialic acid dan isoelectric focusing menunjukkan bahwa seeding density tidak mempengaruhi karakteristik EPO secara signifikan. Namun, apabila dilihat berdasarkan yield dan aktivitas in-vitro EPO, treat D—seeding density 6×107 sel—menunjukkan hasil yang secara signifikan lebih tinggi dibandingkan perlakuan lainnya. Maka, analisis lanjutan dari semua perlakuan dilakukan untuk mengetahui persentase acidic isoform EPO dan persentase tertinggi ditunjukkan oleh treat B dan treat C dengan seeding density 4×107 dan 5×107 sel, secara berturut-turut. Hasil tersebut menunjukan bahwa acidic EPO terbaik dihasilkan pada rentang jumlah sel akhir di treat B, C, dan D (190.968.750 - 256.250.000 sel)
English Abstract
Recombinant human erythropoietin (EPO) production using Chinese Hamster Ovary (CHO) adherent cells can be optimized by determining the optimum cell seeding density. Seeding density as the initial cell number in culture process determine cell growth and productivity, nutrition availability, glycosylation process, and EPO characteristics. Currently, culture process in EPO production is using split ratio. This can interfere process consistency and end-product quality, raising the importance of determining optimum seeding density. This research is conducted by comparing seeding density of 3×107 (A), 4×107 (B), 5×107 (C), 6×107 (D), dan 7×107 (E) cells. Cell is cultured until production stage using serum free media, then supernatant is harvested and purified until IEX2 chromatography. Later, EPO analysis and characterization from each treat is done. Based on the results, it is shown that the impact of seeding density towards final cell number will influence EPO productivity and characteristics. Sialic acid content and isoelectric focusing results showed that seeding density does not affect EPO characteristics significantly. However, based on EPO yield and in-vitro activity, treat D—seeding density of 6×107 cells—conducted significantly higher result than other treatments. Therefore, further analysis of all treatment is carried out to obtain EPO acidic isoform percentage, where highest percentage is shown by treat B and treat C with the seeding density of 4×107 and 5×107 cells, respectively. These results indicate that the final cell number range between treat B, C, and D (190.968.750 - 256.250.000 cells) can yield a prime amount of acidic EPO
| Item Type: | Thesis (Sarjana) |
|---|---|
| Identification Number: | 052410 |
| Uncontrolled Keywords: | Eritropoietin, sel CHO, seeding density-Erythropoietin, CHO cell, seeding density |
| Divisions: | Fakultas Teknologi Pertanian > Keteknikan Pertanian |
| Depositing User: | soegeng sugeng |
| Date Deposited: | 19 Sep 2024 08:04 |
| Last Modified: | 19 Sep 2024 08:04 |
| URI: | http://repository.ub.ac.id/id/eprint/225857 |
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