From the Cover: Human CD1d-glycolipid tetramers generated by in vitro oxidative refolding chromatography
Abstract
CD1 molecules are specialized in presenting lipids to T lymphocytes, but identification and isolation of CD1-restricted lipidspecific T cells has been hampered by the lack of reliable and sensitive techniques. We here report the construction of CD1d-glycolipid tetramers from fully denatured human CD1d molecules by using the technique of oxidative refolding chromatography. We demonstrate that chaperone- and foldase-assisted refolding of denatured CD1d molecules and β2-microglobulin in the presence of synthetic lipids is a rapid method for the generation of functional and specific CD1d tetramers, which unlike previously published protocols ensures isolation of CD1d tetramers loaded with a single lipid species. The use of human CD1d-α-galactosylceramide tetramers for ex vivo staining of peripheral blood lymphocytes and intrahepatic T cells from patients with viral liver cirrhosis allowed for the first time simultaneous analysis of frequency and specificity of natural killer T cells in human clinical samples. Application of this protocol to other members of the CD1 family will provide powerful tools to investigate lipid-specific T cell immune responses in health and in disease.
- Publication:
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Proceedings of the National Academy of Science
- Pub Date:
- March 2001
- DOI:
- 10.1073/pnas.051604498
- Bibcode:
- 2001PNAS...98.3294K
- Keywords:
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- From the Cover