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Pyrogen testing of lipid-based TPN using Mono Mac 6 monocyte cell line and DELFIA
L Moesby1, E W Hansen, J D Christensen
1The Royal Danish School of Pharmacy, Department of Biological Sciences, 2 LUniversitetsparken, 2100 Copenhagen, Denmark.
Objective:
Measurement of lipopolysaccharide (LPS) induced interleukin-6 (IL-6) secretion in Mono Mac 6 cells.
Method:
Dissociation-enhanced lanthanide fluoro immunoassay (DELFIA), a technique based on time-resolved fluorescence.
Results:
A comparison of DELFIA and the B9 bioassay showed a correlation between the results of the two assays. DELFIA did not show any crossreactivity with the human cytokines TNFalpha, IL-1beta, IL-1alpha, IL-2, IL-4, IFNgamma, murine IL-6 or LPS. In aqueous solution the detection limit was 2.5 pg LPS/ml, and in a 25% dilution of a lipid-based total parenteral nutrition (TPN) product it was 25 pg LPS/ml. TPN did not interfere with the DELFIA assay, but the IL-6 secretion from the Mono Mac 6 cells was inhibited at TPN concentrations above 25%.
Conclusion:
A combination of Mono Mac 6 cells with DELFIA was found to be a reliable method for detecting LPS, but further validation by independent laboratories is justified.
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