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S. Tris-tricine and I. , Peptides were separated in high-resolution

. Tris-tricine, Urea SDS-PAGE gels consisting of three sequentially polymerized layers: (i) resolving gel 20% containing 6 M urea; (ii) spacer gel 11% with 4 M urea and (iii) stacking gel 4% 18 . Gels migrated at room temperature in a Mini-PROTEAN ® Tetra Vertical Electrophoresis Cell

, Bio-Rad) and hybridized against streptavidin peroxidase (HRP) conjugate (for FSL-1-biotin) or directly analyzed by fluorescent detection (for FSL-1-fluorescein). SuperSignal ? West Femto Maximum Sensitivity Substrate Western blotting detection reagent (Thermo Scientific) was used as substrate. Chemiluminescent and fluorescent signals were detected with a Chemidoc MP gel imaging system, FSL-1-biotin or FSL-1-fluorescein bands

, Monitoring product formation in 96-well plate format by fluorescence spectroscopy

, Click chemistry samples containing the biotin-conjugated peptide FSL-1 (20 ?L) were diluted in PBST1 (80 ?L) and added to each well of a 96-well plate. The plate was incubated at room temperature while shaking for 1 h in the dark to bind biotin-conjugates onto the streptavidin-coated surface, after which the wells were rinsed six times with PBST2 (PBS with 1% Tween-20), three times with PBST1 (0.05% Tween20) and three times with PBS, Home-made streptavidin-coated microplate were obtained by incubation of 100 ?L of 10 ?g/mL streptavidin per well on 96-well plates (Greiner)

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