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Biến nạp tạm thời trong nuôi cấy tế bào HEK 293 không có huyết tương để sản xuất hiệu quả rFVIII người
Tóm tắt
Bệnh máu khó đông type A là một rối loạn chảy máu do thiếu hụt yếu tố đông máu VIII. Yếu tố VIII tái tổ hợp (rFVIII) là một phương pháp thay thế cho FVIII nguồn huyết tương trong điều trị bệnh máu khó đông type A. Tuy nhiên, việc sản xuất thương mại rFVIII gặp nhiều khó khăn và tốn kém, dẫn đến giá cả cao và sự thiếu hụt sản phẩm, ngay cả ở những quốc gia có nền kinh tế phát triển. Tình hình này có thể được cải thiện bằng cách áp dụng các phương pháp sản xuất hiệu quả hơn. Ở đây, chúng tôi đã đánh giá tiềm năng của việc biến nạp tạm thời trong việc sản xuất rFVIII trong các nuôi cấy tế bào HEK 293 không có huyết tương và điều tra tác động của các nồng độ DNA khác nhau (0,4, 0,6 và 0,8 μg/106 tế bào) và các quá trình biến nạp lặp lại được thực hiện ở 34° và 37°C. Chúng tôi nhận thấy sự giảm trưởng tế bào khi sử dụng nồng độ DNA cao, nhưng không có sự khác biệt đáng kể nào về hiệu suất biến nạp và hoạt tính sinh học của rFVIII. Điều kiện tốt nhất để sản xuất rFVIII đạt được qua việc biến nạp lặp lại ở 34°C với 0,4 μg DNA/106 tế bào, qua đó gần 50 IU rFVIII hoạt tính được sản xuất sau sáu ngày kể từ khi biến nạp. Do đó, việc biến nạp tạm thời trong môi trường nuôi cấy không có huyết tương là một lựa chọn khả thi cho sản xuất rFVIII với năng suất cao. Công việc đang được tiến hành để tối ưu hóa quy trình này và xác nhận khả năng mở rộng của nó.
Từ khóa
#Bệnh máu khó đông type A #yếu tố VIII tái tổ hợp #nuôi cấy tế bào HEK 293 #biến nạp tạm thời #sản xuất hiệu quảTài liệu tham khảo
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