Journal: Pharmaceutics

Article Title: Intestinal Transcytosis of a Protein Cargo and Nanoparticles Mediated by a Non-Toxic Form of Pseudomonas aeruginosa Exotoxin A

doi: 10.3390/pharmaceutics13081171

Figure Lengend Snippet: Isolation and purity assessment of ntPE-TEV-H 6 after E. coli expression. ( a ) Linear diagram of ntPE-TEV-H 6 . ( b ) Enrichment of ntPE-TEV-H 6 using HisTRAP chromatography showing column flow-through (blue arrow, (1) and imidazole-induced elution (red arrow, (2) monitored at 280 nm. ( c ) SDS-PAGE analysis: lane 1, protein from E. coli before the IPTG induction; lane 2, proteins from E. coli after IPTC induction; lane 3, protein extract. ( d ) Size exclusion chromatography of ntPE-TEV-H 6 following HisTRAP chromatography demonstrating probable aggregated (yellow arrow, (3), monomeric (red arrow, (4), and degraded (blue arrow, (5) forms of the protein. ( e ) SDS-PAGE analysis: lanes 1–6 corresponding to collected protein extracts A7–A12, respectively. Material shown in lanes 2–5 was combined and used for conjugation to nanoparticles.

Article Snippet: The ntPE-TEV-H6 protein was captured using a 5 mL prepacked HisTrap HP column (GE Healthcare, Abingdon, UK) and eluted using a linear 20–500 mM gradient of imidazole (Acros Organics, NJ, USA) in a 50 mM Tris pH 8.0 buffer containing 300 mM sodium chloride.

Techniques: Isolation, Expressing, Chromatography, SDS Page, Size-exclusion Chromatography, Conjugation Assay