title: Measurement of poly(ethylene glycol) by cell-based anti-poly(ethylene glycol) ELISA.
abstract: Poly(ethylene glycol) (PEG) is increasingly used in clinical and experimental medicine. However, quantification of PEG and PEGylated small molecules remains laborious and unsatisfactory. In this report, we stably expressed a functional anti-PEG antibody on the surface of BALB 3T3 cells (3T3/alphaPEG cells) to develop a competitive enzyme-linked immunosorbent assay (ELISA) for PEG quantification. The alphaPEG cell-coated plate bound biotinylated PEG(5K) (CH(3)-PEG(5K)-biotin) and CH(3)-PEG(5K)-(131)I more effectively than did a traditional anti-PEG antibody-coated plate. Competitive binding between PEG (2, 5, 10, or 20 kDa) and a known amount of CH(3)-PEG(5K)-biotin allowed construction of a reproducible competition curve. The alphaPEG cell-based competition ELISA measured small molecules derivatized by PEG(2K), PEG(5K), PEG(10K), PEG(20K), and PEG(5K) at concentrations as low as 58.6, 14.6, 3.7, 3.7, and 14.6 ng/mL, respectively. Notably, the presence of serum or bovine serum albumin enhanced PEG measurement by the alphaPEG cell-based competition ELISA. Finally, we show here that the alphaPEG cell-based competition ELISA accurately delineated the pharmacokinetics of PEG(5K), comparable to those determined by direct measurement of radioactivity in blood after intravenous injection of CH(3)-PEG(5K)-(131)I into mice. This quantitative strategy may provide a simple and sensitive method for quantifying PEG and PEGylated small molecules in vivo.
authors.affiliation: Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: Kuo-Hsiang
authors.initials: KH
authors.lastname: Chuang
authors.forename: Shey-Cherng
authors.initials: SC
authors.lastname: Tzou
authors.forename: Ta-Chun
authors.initials: TC
authors.lastname: Cheng
authors.forename: Chien-Han
authors.initials: CH
authors.lastname: Kao
authors.forename: Wei-Lung
authors.initials: WL
authors.lastname: Tseng
authors.forename: Jentaie
authors.initials: J
authors.lastname: Shiea
authors.forename: Kuang-Wen
authors.initials: KW
authors.lastname: Liao
authors.forename: Yun-Ming
authors.initials: YM
authors.lastname: Wang
authors.forename: Ya-Chen
authors.initials: YC
authors.lastname: Chang
authors.forename: Bo-Jyun
authors.initials: BJ
authors.lastname: Huang
authors.forename: Chang-Jer
authors.initials: CJ
authors.lastname: Wu
authors.forename: Pei-Yu
authors.initials: PY
authors.lastname: Chu
authors.forename: Steve R
authors.initials: SR
authors.lastname: Roffler
authors.forename: Tian-Lu
authors.initials: TL
authors.lastname: Cheng
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology*; D041702:BALB 3T3 Cells; D002417:Cattle; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D051379:Mice; D011092:Polyethylene Glycols / Q000032:analysis* / Q000493:pharmacokinetics*; D044967:Serum / Q000737:chemistry; D012710:Serum Albumin, Bovine / Q000737:chemistry
chemical_list: D000906:Antibodies; D012710:Serum Albumin, Bovine; D011092:Polyethylene Glycols