Sensitive quantification of PEGylated compounds by second-generation anti-poly(ethylene glycol) monoclonal antibodies.
Poly(ethylene glycol) (PEG) is often attached to compounds to increase serum half-life, reduce immunogenicity, and enhance bioavailability. Accurate and sensitive quantification of PEG conjugates is critical for product development, pharmacokinetic measurements, and efficacy studies. However, PEGylated compounds can be difficult to quantify due to epitope masking by PEG. We previously generated two monoclonal antibodies to PEG (AGP3, IgM and E11, IgG) for quantitative detection of PEGylated proteins. We now report the identification of two second-generation mAbs to PEG (AGP4, IgM and 3.3, IgG) that bind to the repeating subunits of the PEG backbone and facilitate more sensitive quantification of a wider range of PEGylated compounds. A sandwich ELISA in which AGP4/3.3-biotin was employed as the capture/detection antibodies allowed quantification of PEG-Qdot 525 with 14-50-fold greater sensitivity than the original AGP3/E11 combination. Pegasys (PEG-interferon alpha-2a), PEG-Intron (PEG-interferon alpha-2b), Neulasta (PEG-G-CSF), and Lipo-Dox (PEGylated liposomal doxorubicin) could also be quantified with low ng/mL detection limits. The assay tolerated the presence of 50% human serum or 20% free PEG molecules. These new anti-PEG antibodies appear useful for qualitative and quantitative analysis of a wide range of PEGylated compounds.
Su
Yu-Cheng
YC
Institute of Microbiology and Immunology, National Yang-Ming University, Taipei, 11529, Taiwan.
Chen
Bing-Mae
BM
Chuang
Kuo-Hsiang
KH
Cheng
Tian-Lu
TL
Roffler
Steve R
SR
D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000276:immunology*; D000937:Antigen-Antibody Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D051379:Mice; D008807:Mice, Inbred BALB C; D011092:Polyethylene Glycols / Q000032:analysis* / Q000737:chemistry; D011506:Proteins / Q000032:analysis* / Q000737:chemistry* / Q000276:immunology
D000911:Antibodies, Monoclonal; D011506:Proteins; D011092:Polyethylene Glycols