Functional monoclonal antibodies to p75 neurotrophin receptor raised in knockout mice

J Neurosci Methods. 2006 Nov 15;158(1):109-20. doi: 10.1016/j.jneumeth.2006.05.022. Epub 2006 Jul 7.

Abstract

In this study, p75NTREXONIII knockout mice were used as immune-naive hosts to produce functional antibodies to human p75NTR. Three monoclonal antibodies were produced and named MLR1, MLR2 and MLR3, and isotyped as IgG1, IgG2a and IgG2a, respectively. MLR1 and MLR2 bound to human p75NTR with higher affinity than the well-characterized ME20.4 in ELISA and also recognized p75NTR present on neurons in both rat and mouse. MLR1 and MLR2 bound to nerves known to express p75NTR following injection into Balb/C mice but not p75NTREXONIII knockout mice, indicating the antibodies are directed against the ligand binding extracellular region absent in knockout mice. Both MLR1 and MLR2 partially blocked NGF induced cell death in a mouse cell-line that expresses p75NTR but not TrKA. Importantly, intracerebroventricular injections indicated MLR2 was internalized within the cell bodies of mouse basal forebrain neurons, further demonstrating that this antibody is biologically active.

MeSH terms

  • Animals
  • Antibodies, Monoclonal / physiology*
  • Antibody Specificity*
  • Antigen-Antibody Reactions
  • Blood Vessels / metabolism
  • Blotting, Western / methods
  • Cells, Cultured
  • Enzyme-Linked Immunosorbent Assay
  • Female
  • Flow Cytometry / methods
  • Humans
  • Immunohistochemistry / methods
  • Male
  • Mice
  • Mice, Inbred BALB C
  • Mice, Knockout / immunology*
  • Rats
  • Rats, Sprague-Dawley
  • Receptor, Nerve Growth Factor / genetics*
  • Receptor, Nerve Growth Factor / immunology*

Substances

  • Antibodies, Monoclonal
  • Receptor, Nerve Growth Factor