초록
본 연구는 재조합 단백질 A를 생산하기 위한 정제공정 즉, 생산공정은 양이온교환칼럼(SP)과 음이온교환칼럼(Q)을 통하여 정제한 후 최종 재조합 단백질 A를 생산하게 되는데, 공정단계별로 생산된 시료를 바로 다음단계 공정으로 가지 못하는 경우 보관을 하는데 있어 안정성 확보 문제를 가지고 있다. 이에 본 연구에서 생산공정에서 생산된 시료를 보관 했을때 적정 보관조건과 시간을 알아보고자 하는데 목적이 있다. 즉, 공정시료의 저장방법 및 저장기간 설정과 경시변화에 따른 품질의 안정성의 평가를 목적으로 한다. 실험항목은 엔도톡신, SDS PAGE, HPLC 순도 그리고 농도이다. 실험결과 양이온교환칼럼 통과 후 냉장($4^{\circ}C$), 실온보관에서 SDS PAGE는 major band, 엔도톡신은 5.0 Eu/mg이하, 농도에서는 평균 8.21~8.24 mg/mL, RSD% 0.10~0.62%, 그리고 HPLC 순도 214 nm에서 평균 99.24~99.37%, RSD% 0.22~0.29%, 280 nm에서 평균 89.72~89.80%, RSD% 0.62~1.26%로서 다소 안정된 결과를 나타내었다. 음이온칼럼통과 후 냉장($4^{\circ}C$), 실온보관에서 SDS PAGE major band, 엔도톡신 0.5 Eu/mg이하, 농도는 평균 5.59 mg/mL, RSD% 0.03~0.10% 그리고 HPLC 순도 214 nm에서 평균 99.74%, RSD% 0.10~0.11%, 280 nm에서 평균 96.16~96.85%, RSD% 0.72~1.13%의 결과를 보였다. 상기의 실험결과로 재조합 단백질 A 제조공정시료의 실온과 $4^{\circ}C$에서 7~8 일 그리고 20~21 일 보관은 물질분해 특성이 없이 안정성이 확보 되었다는 결론을 얻을 수 있었다.
This study is to investigate the issues on how to secure stability during the purification process for the production of recombinant protein A. The final recombinant protein A is produced by passing through the cation exchange column (SP) and the anion-exchange column (Q) during the production process, for which the samples produced by the step-by-step processes can be exposed to trouble in securing stable storage in case the next process cannot be taken within the proper time period. Accordingly, this study aims to evaluate the proper storage conditions and length of time when storing samples produced in the production process. That is, in this study, how to store fair samples, how long the storage period should be set up, and how to evaluate the security of its quality depending on time are dealt with. The items to be experimented with were enodotoxin, SDS-PAGE, HPLC purity and concentration. Experimental results showed that after passing the cation exchange column, when stored at $4^{\circ}C$ or room temperature, SDS-PAGE showed a major band, endotoxin is 5.0 Eu/mg or less, and concentration is on average of 8.21 to 8.24 mg/mL and RSD% 0.10~0.62%. In addition, HLPC purity showed somewhat stable results; at the HPLC purity 214 nm, the average is 99.24% to 99.37% and RSD% is 0.22~0.29%, while the average is 89.72% to 89.80% and RSD% 0.62~1.26% at 280 nm. On the contrary, after passing the anion exchange column, when stored at $4^{\circ}C$ or room temperature, SDS-PAGE revealed the major band, endotoxin is 0.5 Eu/mg or less, and concentration is on average of 5.59 mg/mL and RSD% 0.03~0.10%. when it comes to HLPC purity, the result showed that at the HPLC purity 214 nm, the average is 99.74% and RSD% is 0.10~0.11%, while the average is 96.16% to 96.85% and RSD% 0.72~1.13%. In conclusion, the stability of fair samples of recombinant protein A during the manufacturing process could be obtained without substance decomposition for 7~8 days at $4^{\circ}C$ or 20~21 days at room temperature.
