title: ELISA to evaluate plasma anti-asparaginase IgG concentrations in patients with acute lymphoblastic leukemia.
abstract: The development of antibodies to asparaginase may attenuate the pharmacologic effect of asparaginase treatment, may be associated with hypersensitivity reactions, and may necessitate switching to a different commercial asparaginase preparation for current or future therapy. Thus, development of an ELISA for measurement of anti-asparaginase antibody levels is important in the clinical setting. An anti-asparaginase antibody reference was established by screening 65 plasma samples from six patients with acute lymphoblastic leukemia (ALL) who had recently developed a hypersensitivity reaction to Escherichia coli or Erwinia chrysanthemi asparaginase therapy. Twenty-one plasma samples were selected for the anti-asparaginase antibody reference pool. Five micrograms per milliliter of commercial E. coli and Erwinia asparaginase and 10 microg/ml of E. coli asparaginase conjugated with polyethylene glycol (PEG asparaginase) were found to be optimal as coating antigen concentrations. Anti-asparaginase antibody concentrations were determined using a commercial polyclonal goat anti-human IgG horseradish peroxidase conjugate. The antibody reference curves were linear in a range of absorbance from 0.1 to 1. 5 O.D. units for dilutions from 1:1600 to 1:51,200. Inter-assay coefficients of variation were 9.04, 14.7 and 13.0%, and intra-assay coefficients of variation were 1.44, 4.43 and 3.28% for antibodies against E. coli, Erwinia, and PEG L-asparaginase, respectively. The cut-off for positivity in plasma was determined as mean+2 S.D. of the optical density values for plasma from untreated healthy volunteers. Measurement of specific IgG by this ELISA allows for the evaluation of plasma anti-asparaginase antibody concentrations in patients receiving one or more of the multiple commercial L-asparaginase preparations.
authors.affiliation: Department of Clinical Pharmacy and the Center for Pediatric Pharmacokinetics and Therapeutics, College of Pharmacy, University of Tennessee, 26 South Dunlap Street, Memphis, TN, USA. bwang@utmem.edu
authors.forename: B
authors.initials: B
authors.lastname: Wang
authors.forename: L J
authors.initials: LJ
authors.lastname: Hak
authors.forename: M V
authors.initials: MV
authors.lastname: Relling
authors.forename: C H
authors.initials: CH
authors.lastname: Pui
authors.forename: M H
authors.initials: MH
authors.lastname: Woo
authors.forename: M C
authors.initials: MC
authors.lastname: Storm
mesh_terms: D000042:Absorption; D001215:Asparaginase / Q000276:immunology* / Q000627:therapeutic use; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000592:standards; D004885:Erwinia / Q000201:enzymology; D004926:Escherichia coli / Q000201:enzymology; D006735:Horseradish Peroxidase; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood*; D007202:Indicators and Reagents; D016014:Linear Models; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000188:drug therapy / Q000201:enzymology / Q000276:immunology*; D015203:Reproducibility of Results
chemical_list: D007074:Immunoglobulin G; D007202:Indicators and Reagents; D006735:Horseradish Peroxidase; D001215:Asparaginase