Shabani Mahdi, Hemmati Sheyda, Hadavi Reza, Amirghofran Zahra, Jeddi-Tehrani Mahmood, Rabbani Hodjatallah, Shokri Fazel
Department of Immunology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran ; Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.
Avicenna J Med Biotechnol. 2010 Jul;2(3):123-30.
Purification and isolation of cellular target proteins for monoclonal antibody (MAb) production is a difficult and time-consuming process. Immunization of mice with murine cell lines stably transfected with genes coding for xenogenic target molecules is an alternative method for mouse immunization and MAb production. Here we present data on transfection efficiency of some commercial reagents used for transfection of murine myeloma cell lines. Little is known about transfectability of murine myeloma cell lines by different transfection reagents. Mouse myeloma cell lines (SP2/0, NS0, NS1, Ag8, and P3U1) were transfected with pEGFP-N1 vector using Lipofectamine 2000, jetPEI and LyoVec commercial transfection reagents in different combinations. The transfection permissible HEK293-FT cell line was used as a control in transfection procedure. Transfected cells, expressing the Enhanced Green Fluorescent Protein (EGFP), were analyzed by flow cytometry 48 hrs post transfection. Our results showed transfection efficiency of 71%, 57% and 22% for HEK293-FT, 5.5%, 3.4% and 1% for SP2/0, 55.7%, 21.1% and 9.3% for NS0, 8.2%, 6% and 5.5% for NS1, 22%, 49.2% and 5.5% for Ag8 and 6.3%, 21.5% and 4.6% for P3U1 cell lines after transfection with Lipofectamine 2000, jetPEI and LyoVec reagents, respectively. Our data indicate that NS0 and Ag8 are efficiently transfected by Lipofectamine 2000 and jetPEI reagents. Finally, we propose Ag8 and NS0 cell lines as suitable host cells for efficient expression of target genes which can be used for mouse immunization and MAb production.
用于单克隆抗体（MAb）生产的细胞靶蛋白的纯化和分离是一个困难且耗时的过程。用稳定转染了编码异种靶分子基因的鼠细胞系免疫小鼠是小鼠免疫和单克隆抗体制备的一种替代方法。在此，我们展示了一些用于转染鼠骨髓瘤细胞系的商业试剂的转染效率数据。关于不同转染试剂对鼠骨髓瘤细胞系的转染能力了解甚少。使用Lipofectamine 2000、jetPEI和LyoVec商业转染试剂，以不同组合将鼠骨髓瘤细胞系（SP2/0、NS0、NS1、Ag8和P3U1）用pEGFP-N1载体进行转染。转染允许的HEK293-FT细胞系用作转染过程的对照。转染后48小时，通过流式细胞术分析表达增强型绿色荧光蛋白（EGFP）的转染细胞。我们的结果显示，用Lipofectamine 2000、jetPEI和LyoVec试剂转染后，HEK293-FT细胞系的转染效率分别为71%、57%和22%，SP2/0细胞系分别为5.5%、3.4%和1%，NS0细胞系分别为55.7%、21.1%和9.3%，NS1细胞系分别为8.2%、6%和5.5%，Ag8细胞系分别为22%、49.2%和5.5%，P3U1细胞系分别为6.3%､21.5%和4.6%。我们的数据表明，NS0和Ag8细胞系能用Lipofectamine 2000和jetPEI试剂有效转染。最后，我们提出Ag8和NS0细胞系作为可用于高效表达靶基因的合适宿主细胞，这些靶基因可用于小鼠免疫和单克隆抗体制备。
