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. 2016 Jul 15;5(9):e1211220. doi: 10.1080/2162402X.2016.1211220

Figure 1.

Figure 1.

Schematic representation and expression of ULBP2-aCD19, ULBP2-aCD33, ULBP2-aCD19-aCD33 and ULBP2-aCD19-aCD19 immunoligands. (A) Cartoon illustrating bispecific immunoligands ULBP2-aCD19 (U-19) and ULBP2-aCD33 (U-33) and triplebodies ULBP2-aCD19-aCD33 (U-19-33) and ULBP2-aCD19-aCD19 (U-19-19) as well as control constructs: aCD19scFv and aCD33scFv. All four “test” immunoligands contain 15mer Gly-Ser linker (GGGGS)3x to provide flexibility and a c-Myc tag and 6xHis tag for purification/detection. An N-terminal part of ULBP2 facilitated secretion of all four immunoligands into the s/n of transfected cells. In contrast, Igκ leader sequence was placed 3′ of both control constructs (aCD19scFv and aCD33scFv) to facilitate their secretion and 6xHis tag was used for purification. (B and C) All constructs were expressed in eukaryotic cell line HEK293T and purified from the supernatant by affinity chromatography utilizing their 6xHis tag. All four (U-19, U-33, U-19-33 and U-19-19) immunoligands were separated on SDS-PAGE for protein gel blot staining (B) using anti c-Myc tag antibody or coomassie staining (C) to confirm the size and purity. Coomassie staining was also done to check the size and purity of both control constructs aCD19scFv and aCD33scFv (Data not shown).