PMID- 21827863
OWN - NLM
STAT- MEDLINE
DA  - 20111125
DCOM- 20120608
IS  - 1873-488X (Electronic)
IS  - 1056-8719 (Linking)
VI  - 64
IP  - 3
DP  - 2011 Nov-Dec
TI  - A double antigen bridging immunogenicity ELISA for the detection of antibodies to
      polyethylene glycol polymers.
PG  - 238-45
LID - 10.1016/j.vascn.2011.07.003 [doi]
AB  - INTRODUCTION: Polyethylene glycol (PEG) polymers attached to biotherapeutic
      molecules enhance in vivo delivery and stability of these large molecular weight 
      drugs. However, these polymers may by themselves be immunogenic and elicit
      antibodies that can reduce the efficacy of the drug and contribute to potential
      patient morbidity. A double antigen bridging ELISA immunogenicity assay for the
      detection of anti-drug antibodies (ADAs) specific to PEG polymers of various
      sizes has been developed. METHODS: Hapten-labeled conjugate of 40kDa PEG polymer 
      was synthesized and used in a double antigen bridging ELISA. The hapten-labeled
      PEG is incubated with the patient sample, then this mixture is added to a 96-well
      microplate precoated with 40kDa PEG, allowing PEG-specific ADA to form a bridge
      complex with the PEG conjugate and the PEG coated on the microplate. After
      incubation, the reaction mixture is removed and replaced by horseradish
      peroxidase (HRP)-labeled anti-hapten antibody. After sufficient incubation, the
      plate is washed and substrate reagent is added. Enzyme color development,
      directly proportional to ADA, is stopped after 20min with 2N sulfuric acid and
      the absorbance in each well is measured at 450/630nm. Dose response, drug
      tolerance, matrix effects, reproducibility, specificity/free drug depletion
      experiments and screening cut-point determination of 350 naive normal human sera 
      were performed. RESULTS: Using an anti-PEG mouse monoclonal IgM as a positive
      control, a reproducible dose response curve was demonstrated for the PEG
      Immunogenicity ELISA. Pre-existing PEG-specific antibodies which were proven to
      be highly specific to the PEG polymer structure were found in 15 human serum
      samples in a total population of 350 naive donors. The assay exhibited no
      significant matrix effects and was shown to be highly reproducible. DISCUSSION: A
      double antigen bridging immunogenicity assay for the detection of antibodies to
      PEG in the typical polymer size ranges used in biotherapeutics has been
      successfully developed in ELISA format. The antibodies detected in positive
      samples displayed a diverse spectrum of specificities for different PEG polymer
      lengths and linking functional groups. The discovery of 15 confirmed positive
      samples among 350 naive patient samples calls into focus the need for testing
      PEG-specific immunogenicity of PEGylated biotherapeutics.
CI  - Copyright (c) 2011 Elsevier Inc. All rights reserved.
AD  - ANP Technologies, Inc. 824 Interchange Blvd., Newark, DE 19711, USA.
FAU - Liu, Yijuan
AU  - Liu Y
FAU - Reidler, Helen
AU  - Reidler H
FAU - Pan, Jing
AU  - Pan J
FAU - Milunic, David
AU  - Milunic D
FAU - Qin, Dujie
AU  - Qin D
FAU - Chen, Dave
AU  - Chen D
FAU - Vallejo, Yli Remo
AU  - Vallejo YR
FAU - Yin, Ray
AU  - Yin R
LA  - eng
PT  - Journal Article
DEP - 20110729
PL  - United States
TA  - J Pharmacol Toxicol Methods
JT  - Journal of pharmacological and toxicological methods
JID - 9206091
RN  - 0 (Antibodies)
RN  - 0 (Antibodies, Anti-Idiotypic)
RN  - 0 (Antigens)
RN  - 0 (Drug Carriers)
RN  - 0 (Haptens)
RN  - 0 (Polyethylene Glycols)
RN  - 9002-88-4 (Polyethylene)
SB  - IM
MH  - Antibodies/*analysis/immunology
MH  - Antibodies, Anti-Idiotypic/immunology
MH  - Antigens/*immunology
MH  - Drug Carriers/pharmacology
MH  - Drug Tolerance/immunology
MH  - Enzyme-Linked Immunosorbent Assay/*methods
MH  - Haptens/immunology
MH  - Humans
MH  - Polyethylene/*immunology
MH  - Polyethylene Glycols/*pharmacology
MH  - Reproducibility of Results
MH  - Sensitivity and Specificity
MH  - Serum/immunology
EDAT- 2011/08/11 06:00
MHDA- 2012/06/09 06:00
CRDT- 2011/08/11 06:00
PHST- 2011/03/23 [received]
PHST- 2011/07/15 [revised]
PHST- 2011/07/18 [accepted]
PHST- 2011/07/29 [aheadofprint]
AID - S1056-8719(11)00269-3 [pii]
AID - 10.1016/j.vascn.2011.07.003 [doi]
PST - ppublish
SO  - J Pharmacol Toxicol Methods. 2011 Nov-Dec;64(3):238-45. doi:
      10.1016/j.vascn.2011.07.003. Epub 2011 Jul 29.