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Immunoaffinity LC–MS/MS is more suitable than ELISA to quantify a PEGylated molecule in cynomolgus monkey serum

by Dai, Lu , Kaur, Surinder , Liu, Yanqiu , Stella, Cinzia , Liu, Luna , Yu, Nancy , Chang, Vincent , Wakshull, Eric , Xu, Keyang

in Antibodies / Atrophy / Binding sites / Drug dosages / Edema / ELISA / Enzyme-linked immunosorbent assay / IA LC–MS/MS / immunoaffinity LC–MS/MS / intravitreal / ITV / LAD / Liquid chromatography / long acting delivery / Macular degeneration / Mass spectroscopy / PEGylation / Peptides / Photoreceptors / Polyethylene / Polyethylene glycol / Proteins / Quantitation / Reference materials

2020

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Immunoaffinity LC–MS/MS is more suitable than ELISA to quantify a PEGylated molecule in cynomolgus monkey serum

by Dai, Lu , Kaur, Surinder , Liu, Yanqiu , Stella, Cinzia , Liu, Luna , Yu, Nancy , Chang, Vincent , Wakshull, Eric , Xu, Keyang

2020

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Immunoaffinity LC–MS/MS is more suitable than ELISA to quantify a PEGylated molecule in cynomolgus monkey serum

by Dai, Lu , Kaur, Surinder , Liu, Yanqiu , Stella, Cinzia , Liu, Luna , Yu, Nancy , Chang, Vincent , Wakshull, Eric , Xu, Keyang

2020

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Journal Article

Immunoaffinity LC–MS/MS is more suitable than ELISA to quantify a PEGylated molecule in cynomolgus monkey serum

Dai, Lu,

Kaur, Surinder,

Liu, Yanqiu,

Stella, Cinzia,

Liu, Luna,

Yu, Nancy,

Chang, Vincent,

Wakshull, Eric,

Xu, Keyang

2020

Overview

Polyethylene glycolylation (PEGylation) technology is a long-acting delivery platform used to increase the half-life of protein therapeutics. Quantitation of PEGylated anti-Factor D Fab (PEG-aFD) poses bioanalytical challenges. An ELISA was developed to determine total Fab concentration in cynomolgus monkey serum following intravitreal administration of PEG-aFD. However, assay characterization showed a low recovery of about 25% for free unconjugated Fab whereas recovery for PEG-conjugated Fab was within 80–120%. To overcome this challenge, an immunoaffinity liquid chromatography tandem mass spectrometry (IA LC–MS/MS) assay was developed, achieving recovery within 80–120% for both free and conjugated Fab. Immunoaffinity LC–MS/MS is more suitable than ELISA to accurately quantify the total protein concentration of PEG-aFD in cynomolgus monkey serum.

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Publisher

Newlands Press Ltd,Newlands Press

Subject

Antibodies

/ Atrophy

/ Binding sites

/ Drug dosages

/ Edema

/ ELISA

/ Enzyme-linked immunosorbent assay