Molecular Biotechnology

, Volume 46, Issue 3, pp 279–286

Generation of Polyclonal Antibodies Against Recombinant Human Glucocerebrosidase Produced in Escherichia coli

  • Juliana Branco Novo
  • Maria Leonor Sarno Oliveira
  • Geraldo Santana Magalhães
  • Ligia Morganti
  • Isaías Raw
  • Paulo Lee Ho
Research

DOI: 10.1007/s12033-010-9303-4

Cite this article as:
Novo, J.B., Oliveira, M.L.S., Magalhães, G.S. et al. Mol Biotechnol (2010) 46: 279. doi:10.1007/s12033-010-9303-4

Abstract

Deficiency of the lysosomal glucocerebrosidase (GCR) enzyme results in Gaucher’s disease, the most common inherited storage disorder. Treatment consists of enzyme replacement therapy by the administration of recombinant GCR produced in Chinese hamster ovary cells. The production of anti-GCR antibodies has already been described with placenta-derived human GCR that requires successive chromatographic procedures. Here, we report a practical and efficient method to obtain anti-GCR polyclonal antibodies against recombinant GCR produced in Escherichia coli and further purified by a single step through nickel affinity chromatography. The purified GCR was used to immunize BALB/c mice and the induction of anti-GCR antibodies was evaluated by enzyme-linked immunosorbent assay. The specificity of the antiserum was also evaluated by western blot analysis against recombinant GCR produced by COS-7 cells or against endogenous GCR of human cell lines. GCR was strongly recognized by the produced antibodies, either as cell-associated or as secreted forms. The detected molecular masses of 59–66 kDa are in accordance to the expected size for glycosylated GCR. The GCR produced in E. coli would facilitate the production of polyclonal (shown here) and monoclonal antibodies and their use in the characterization of new biosimilar recombinant GCRs coming in the near future.

Keywords

Glucocerebrosidase Polyclonal antibody Gaucher’s disease Recombinant protein expression Lysosomal storage disorder 

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Juliana Branco Novo
    • 1
    • 2
  • Maria Leonor Sarno Oliveira
    • 1
  • Geraldo Santana Magalhães
    • 3
  • Ligia Morganti
    • 4
  • Isaías Raw
    • 1
  • Paulo Lee Ho
    • 1
    • 2
  1. 1.Centro de Biotecnologia, Instituto ButantanSão PauloBrazil
  2. 2.Interunidades em Biotecnologia, Instituto de Ciências Biomédicas, Universidade de São PauloSão PauloBrazil
  3. 3.Laboratório de Imunopatologia, Instituto ButantanSão PauloBrazil
  4. 4.Centro de Biotecnologia, Instituto de Pesquisas Energéticas e NuclearesSão PauloBrazil

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