A photobleaching energy transfer analysis of CD8/MHC-I and LFA-1/ICAM-1 interactions in CTL-target cell conjugates

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Abstract

The photobleaching energy transfer (pbFRET) technique is a fluorescence method to measure proximity relationships between molecules, especially cell surface proteins, labeled with fluorophore-conjugated monoclonal antibodies, on a pixel-by-pixel base using digital imaging microscopy. This technique enables analysis of inter- and intramolecular proximities at cell surfaces at physiological conditions. We have developed a pbFRET approach to measure intercellular proximities in order to access spatial organization of interacting proteins in the contact region of two 'communicating' cells. Two examples, as possible application areas of this approach, are presented here: interaction between CD8 and MHC-I molecules in point contacts and interaction between LFA-1 and ICAM-1 molecules in focal contacts of CTL-target conjugates. The geometry of these protein contacts based on our resonance energy transfer (RET) data is consistent with the observed blocking effects of monoclonal antibodies (directed against the interacting proteins) on the cytolytic activity of CTLs and suggest a critical role for CD8β-subunit in signal transmission in peripheral T-lymphocytes.

Original languageEnglish
Pages (from-to)151-156
Number of pages6
JournalImmunology Letters
Volume54
Issue number2-3
DOIs
Publication statusPublished - Dec 2 1996

Fingerprint

Photobleaching
Lymphocyte Function-Associated Antigen-1
Energy Transfer
Intercellular Adhesion Molecule-1
Monoclonal Antibodies
Proteins
Focal Adhesions
Microscopy
Membrane Proteins
Fluorescence
T-Lymphocytes

Keywords

  • antigen recognition
  • cytotoxic CD8 T-cells
  • europium-release
  • ICAM-1
  • JY cells
  • LFA-1
  • MHC-I
  • photobleaching energy transfer

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy

Cite this

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title = "A photobleaching energy transfer analysis of CD8/MHC-I and LFA-1/ICAM-1 interactions in CTL-target cell conjugates",
abstract = "The photobleaching energy transfer (pbFRET) technique is a fluorescence method to measure proximity relationships between molecules, especially cell surface proteins, labeled with fluorophore-conjugated monoclonal antibodies, on a pixel-by-pixel base using digital imaging microscopy. This technique enables analysis of inter- and intramolecular proximities at cell surfaces at physiological conditions. We have developed a pbFRET approach to measure intercellular proximities in order to access spatial organization of interacting proteins in the contact region of two 'communicating' cells. Two examples, as possible application areas of this approach, are presented here: interaction between CD8 and MHC-I molecules in point contacts and interaction between LFA-1 and ICAM-1 molecules in focal contacts of CTL-target conjugates. The geometry of these protein contacts based on our resonance energy transfer (RET) data is consistent with the observed blocking effects of monoclonal antibodies (directed against the interacting proteins) on the cytolytic activity of CTLs and suggest a critical role for CD8β-subunit in signal transmission in peripheral T-lymphocytes.",
keywords = "antigen recognition, cytotoxic CD8 T-cells, europium-release, ICAM-1, JY cells, LFA-1, MHC-I, photobleaching energy transfer",
author = "Z. Bacs{\'o} and L. Bene and Andrea Bodn{\'a}r and J. Matk{\'o} and S. Damjanovich",
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T1 - A photobleaching energy transfer analysis of CD8/MHC-I and LFA-1/ICAM-1 interactions in CTL-target cell conjugates

AU - Bacsó, Z.

AU - Bene, L.

AU - Bodnár, Andrea

AU - Matkó, J.

AU - Damjanovich, S.

PY - 1996/12/2

Y1 - 1996/12/2

N2 - The photobleaching energy transfer (pbFRET) technique is a fluorescence method to measure proximity relationships between molecules, especially cell surface proteins, labeled with fluorophore-conjugated monoclonal antibodies, on a pixel-by-pixel base using digital imaging microscopy. This technique enables analysis of inter- and intramolecular proximities at cell surfaces at physiological conditions. We have developed a pbFRET approach to measure intercellular proximities in order to access spatial organization of interacting proteins in the contact region of two 'communicating' cells. Two examples, as possible application areas of this approach, are presented here: interaction between CD8 and MHC-I molecules in point contacts and interaction between LFA-1 and ICAM-1 molecules in focal contacts of CTL-target conjugates. The geometry of these protein contacts based on our resonance energy transfer (RET) data is consistent with the observed blocking effects of monoclonal antibodies (directed against the interacting proteins) on the cytolytic activity of CTLs and suggest a critical role for CD8β-subunit in signal transmission in peripheral T-lymphocytes.

AB - The photobleaching energy transfer (pbFRET) technique is a fluorescence method to measure proximity relationships between molecules, especially cell surface proteins, labeled with fluorophore-conjugated monoclonal antibodies, on a pixel-by-pixel base using digital imaging microscopy. This technique enables analysis of inter- and intramolecular proximities at cell surfaces at physiological conditions. We have developed a pbFRET approach to measure intercellular proximities in order to access spatial organization of interacting proteins in the contact region of two 'communicating' cells. Two examples, as possible application areas of this approach, are presented here: interaction between CD8 and MHC-I molecules in point contacts and interaction between LFA-1 and ICAM-1 molecules in focal contacts of CTL-target conjugates. The geometry of these protein contacts based on our resonance energy transfer (RET) data is consistent with the observed blocking effects of monoclonal antibodies (directed against the interacting proteins) on the cytolytic activity of CTLs and suggest a critical role for CD8β-subunit in signal transmission in peripheral T-lymphocytes.

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KW - cytotoxic CD8 T-cells

KW - europium-release

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KW - LFA-1

KW - MHC-I

KW - photobleaching energy transfer

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