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Determination of interferon‐gamma in Chinese hamster ovary cell culture supernatant by coupled‐column liquid chromatography
Author(s) -
Goh LinTang,
Yap Miranda Gek Sim
Publication year - 2005
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.200500045
Subject(s) - chinese hamster ovary cell , chromatography , chemistry , high performance liquid chromatography , detection limit , quantitative analysis (chemistry) , resolution (logic) , reversed phase chromatography , biochemistry , receptor , artificial intelligence , computer science
A robust tandem HPLC method coupling size‐exclusion (Shodex Asahipak GS‐320HQ) and reversed phase (Vydac 218TP54) columns with ultraviolet detection was developed for quantitative determination of interferon‐γ (IFN‐γ) in Chinese hamster ovary cell culture supernatant. The 2D‐HPLC system was linked up by a 6‐port 2‐position low hold‐up volume switch valve. Compared to a commercial ELISA kit for IFN‐γ, the coupled column LC approach was able to detect and quantify soluble IFN‐γ, regardless of the glycoprotein's molecular/conformational variability and sample background. Each LC‐LC analysis took 90 minutes inclusive of column regeneration. The relative standard deviation of measurements ( n = 5) was less than 3%. The limit of detection (LOD) was determined to be 0.35 μg IFN‐γ.