HPLC quantification of protein

LB Laura Bryan
MH Michael Henry
NB Niall Barron
CG Clair Gallagher
RK Ronan M. Kelly
CF Christopher C. Frye
MO Matthew D. Osborne
MC Martin Clynes
PM Paula Meleady
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Cell free supernatant media was taken from CHO CDCLs transfected with miRNA mimics or negative control mimics on day 5 of culture and absorbance peaks were obtained. Cell free supernatant was run on an UltiMate 3000 RSLC HPLC using a MAbPacTM RP 4 μm 2.1 × 100 mm column (large pore size polymeric resin) (Thermo Scientific). A flow rate of 500 μL/min, a temperature of 75 °C and an absorbance of 214 nm was applied. The peak obtained represented the amount of full-length product (FLP) present on each day of culture. A standard curve was created using the purified recombinant protein product to determine titer.

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