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Fig. 2 | Microbial Cell Factories

Fig. 2

From: Screening and purification of nanobodies from E. coli culture supernatants using the hemolysin secretion system

Fig. 2

Identification of Nb binders secreted with the Hly-system from a VHH immune library against EHEC antigens. a Coomassie staining of purified EHEC protein antigens EspA, Int280 and TirM used for camel immunization. b ELISA of camel serum after immunization to reveal antibody response against EspA, Int280, TirM and BSA (negative control). The camel antibody response against each of the proteins using the indicated serum dilutions was developed with protein-A peroxidase (POD). c The VHH sequences amplified from the immunized animal were used to generate a phage antibody (Phab) library. Phab binders were enriched by panning to obtain VHH repertoires against each antigen (Ag). The VHH repertoires were cloned into pEHLYA5 and the Nb-HlyA fusions were secreted in E. coli bacteria carrying pVDL9.3. The culture supernatants of individual clones from each VHH repertoire were tested by ELISA against their corresponding antigen (EspA, Int280, TirM) and BSA (negative control). Bound Nb-HlyA fusions were developed with anti-E-tag-mAb

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