Author: Boyington, Jeffrey C.; Joyce, M. Gordon; Sastry, Mallika; Stewart-Jones, Guillaume B. E.; Chen, Man; Kong, Wing-Pui; Ngwuta, Joan O.; Thomas, Paul V.; Tsybovsky, Yaroslav; Yang, Yongping; Zhang, Baoshan; Chen, Lei; Druz, Aliaksandr; Georgiev, Ivelin S.; Ko, Kiyoon; Zhou, Tongqing; Mascola, John R.; Graham, Barney S.; Kwong, Peter D.
Title: Structure-Based Design of Head-Only Fusion Glycoprotein Immunogens for Respiratory Syncytial Virus Document date: 2016_7_27
ID: 1nbocmux_10
Snippet: Antibodies were expressed by transient co-transfection of HEK 293-F cells (Thermo Fisher Scientific, MA) with heavy-and light-chain plasmids using 293fectin (Thermo Fisher Scientific, MA). Cell supernatants were harvested after 4-5 days and passed over Protein A agarose (GE Healthcare, PA). Bound antibodies were washed with PBS and eluted with IgG elution buffer (Pierce, IL) into 1/10th volume of 1 M Tris-HCl pH 8.0. Fabs were created by digestin.....
Document: Antibodies were expressed by transient co-transfection of HEK 293-F cells (Thermo Fisher Scientific, MA) with heavy-and light-chain plasmids using 293fectin (Thermo Fisher Scientific, MA). Cell supernatants were harvested after 4-5 days and passed over Protein A agarose (GE Healthcare, PA). Bound antibodies were washed with PBS and eluted with IgG elution buffer (Pierce, IL) into 1/10th volume of 1 M Tris-HCl pH 8.0. Fabs were created by digesting the IgG with Lys-C or HRV3C protease [24] , and the cleaved Fc region was removed by passing the mixture over Protein A agarose. Fabs were further purified by SEC.
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