Journal of Fluorescence

, Volume 5, Issue 3, pp 273–277 | Cite as

Fluorescence spectroscopy of monoclonal antibodies produced against the fluorescyl hapten conjugated through the xanthene ring

  • P. R. Droupadi
  • T. Nanavaty
  • C. Smith
  • D. D. Johnson
  • M. Adamczyk
  • D. S. Linthicum


Two mouse anti-fluorescyl monoclonal antibodies (mAb), clones FL43.1 and FL55.3, were produced to the fluorescein hapten, which was conjugated to the carrier protein through the 4′ position of the xanthene ring. Association constants (KA) and thermodynamic parameters for both mAb were ascertained by monitoring the steady-state intrinsic and fluorescein fluorescence. Both techniques were in good agreement and gaveKA values in the 109M−1 range. Ligand-induced intrinsic fluorescence quenching showed a hypsochromic shift for mAb FL43.1, but not for FL55.3, suggesting that the ligand interacts with different tryptophan residues in each mAb. Because these mAb are directed toward the phenylcarboxylate portion of fluorescein, the different ionic and structural forms should be useful as indicators of antibody binding site pH and buffering capacity near the binding site.

Key words

Monoclonal antibody fluorescein complex fluorescence quenching intrinsic fluorescence 


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Copyright information

© Plenum Publishing Corporation 1995

Authors and Affiliations

  • P. R. Droupadi
    • 1
  • T. Nanavaty
    • 1
  • C. Smith
    • 1
  • D. D. Johnson
    • 2
  • M. Adamczyk
    • 2
  • D. S. Linthicum
    • 1
  1. 1.Department of Veterinary Pathobiology, College of Veterinary MedicineTexas A&M UniversityCollege Station
  2. 2.Abbott Diagnostics DivisionAbbott LaboratoriesAbbott Park

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