Assessing viability and cell-associated product of recombinant protein producing Pichia pastoris with flow cytometry

被引：105

作者：

Hohenblum, H ^{[1
]}

Borth, N ^{[1
]}

Mattanovich, D ^{[1
]}

机构：

[1] Univ Nat Resources & Appl Life Sci, BOKU, Inst Appl Microbiol, A-1190 Vienna, Austria

来源：

JOURNAL OF BIOTECHNOLOGY | 2003年 / 102卷 / 03期

关键词：

Pichia pastoris; fed batch; heterologous protein; methanol induction; viability; immunofluorescence;

D O I：

10.1016/S0168-1656(03)00049-X

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

This paper describes the establishment of flow cytometric methods for recombinant Pichia pastoris strains, and their application to a lab scale fed batch fermentation. Using a strain which secretes human trypsinogen, the viability and the product which remained associated to the cell were measured with propidium iodide and immunofluorescent staining, respectively. Viability decreases significantly below 70% during the methanol fed batch phase, indicating a stress situation triggered by the fermentation conditions. Cell associated product is accumulated earlier after methanol induction than secreted product. These data demonstrate that flow cytometry is a powerful tool for the analysis and optimization of recombinant protein production processes, and they indicate the need to further improve a widely used fermentation protocol for P. pastoris. (C) 2003 Elsevier Science B.V. All rights reserved.

引用

页码：281 / 290

页数：10