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A Sensitive Enzyme Immunoassay for the Quantitation of Human CTLA4Ig Fusion Protein in Mouse Serum: Pharmacokinetic Application to Optimizing Cell Line Selection

A Sensitive Enzyme Immunoassay for the Quantitation of Human CTLA4Ig Fusion Protein in Mouse Serum: Pharmacokinetic Application to Optimizing Cell Line Selection

R S Weiner, N R Srinivas, J D Calore, C G Fadrowski, W C Shyu, L K Tay

J Pharm Biomed Anal. 1997 Feb;15(5):571-9.

PMID: 9127268

Abstract:

A sensitive, accurate, and precise enzyme immunoassay (EIA) for the quantitation of human CTLA4Ig in mouse serum was validated. The EIA method employed a technique in which a monoclonal anti-CTLA4 antibody was adsorbed onto 96-well polystyrene microtiter plates and used to capture the CTLA4Ig in mouse serum samples. The captured CTLA4Ig was then detected using a goat anti-human IgGFc antiserum conjugated to the enzyme horseradish peroxidase. The validation included assessments of method accuracy and precision, range of reliable response, lower limit of quantitation (LLQ), inter-analyst robustness, storage stability in mouse serum and assay specificity. The results indicate that this validated assay is precise, accurate, and reproducible. This EIA has a range of reliable response in 10% mouse serum of 0.14-4.58 ng ml-1 resulting in a 100% serum equivalent curve of 1.4-45.8 ng ml-1. Assessment of individual standard curve variations indicated a reproducible response with R2 values of > or = 0.995. The LLQ was established at 1.4 ng ml-1. The accuracy and precision estimates, based on the quality control values, were within 3.8% and 5.2% respectively, for CTLA4Ig. Stability of CTLA4Ig was established in mouse serum for 5 days at both 4 degrees C and room temperature, for 2 months at -70 degrees C and through five freeze-thaw cycles. This validated assay was successfully employed in the assessment of pharmacokinetic characteristics of CTLA4Ig in mice and to aid in the selection of an optimal CTLA4Ig-producing cell line.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
IAR42414738	Monoclonal Anti-CTLA4 antibody produced in mouse			Price

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