Teruya K, Shirahata S, Yano T, Seki K, Tachibana H, Ohashi H, Murakami H
Graduate School of Genetic Resources Technology, Faculty of Agriculture, Kyushu University, Fukuoka, Japan.
Anal Biochem. 1993 Nov 1;214(2):468-73. doi: 10.1006/abio.1993.1524.
We have developed a rapid and sensitive screening method to detect the secretory product from transformed secretory cells. Polyclonal antibodies against the target secretory protein are first adsorbed onto a nitrocellulose membrane and are then called solid-phase antibodies. Then this membrane is placed on top of target protein-secreting adherent animal cell colonies. After trapping the secreted protein on the membrane, the protein is detected using the highly sensitive enhanced chemiluminescence detection system. Thus, this method is named the secretory cell immunoscreening assay (SCISA). We have screened recombinant BHK-21 cells, which secrete human interleukin-6 (hIL-6) for highly productive clones. With the SCISA method, it is possible to detect less than 100 pg of hIL-6 secreted by a single colony. We are also able to select highly productive colonies rapidly and easily from a large sample population of transformed cell colonies.
我们开发了一种快速灵敏的筛选方法,用于检测转化分泌细胞的分泌产物。首先将针对目标分泌蛋白的多克隆抗体吸附到硝酸纤维素膜上,这些抗体随后被称为固相抗体。然后将此膜置于分泌目标蛋白的贴壁动物细胞集落之上。在膜上捕获分泌蛋白后,使用高灵敏度的增强化学发光检测系统检测该蛋白。因此,此方法被命名为分泌细胞免疫筛选测定法(SCISA)。我们已对分泌人白细胞介素-6(hIL-6)的重组BHK-21细胞进行筛选,以获得高产克隆。使用SCISA方法,能够检测到单个集落分泌的少于100 pg的hIL-6。我们还能够从大量转化细胞集落样本中快速轻松地筛选出高产集落。