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Differential scanning fluorimetry (DSF)

PS Pietro Sormanni
LA Leanne Amery
SE Sofia Ekizoglou
MV Michele Vendruscolo
BP Bojana Popovic
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Conformational stability was assessed for triplicate samples in 10 mM citrate/phosphate, pH 5.5 by differential scanning fluorimetry. Measurements were conducted on a CFX96 RT-PCR system (BIO-RAD) in a 96-well format combining mAbs 1–9 at 10 mg/mL in 25 mM acetate, 240 mM sucrose, pH 5.0 with 10 mM citrate/phosphate, pH 5.5 and SYPRO orange fluorophor (Invitrogen) to give a final concentration of 1 mg/mL. Samples were heated from 20 °C to 95 °C at 0.2 °C/min. Melt curves were generated automatically in CFX Manager (BIO-RAD) software with fluorescence intensities plotted as a function of temperature and melt temperatures determined.

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