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Detection and characterization of cellular immune responses using peptide-MHC microarrays.

Soen Y, Chen DS, Kraft DL, Davis MM, Brown PO - PLoS Biol. (2003)

Bottom Line: The detection and characterization of antigen-specific T cell populations is critical for understanding the development and physiology of the immune system and its responses in health and disease.We have developed and tested a method that uses arrays of peptide-MHC complexes for the rapid identification, isolation, activation, and characterization of multiple antigen-specific populations of T cells.In addition, we were able to use the array to detect a rare population of antigen-specific T cells following vaccination of a normal mouse.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Stanford University, Stanford, California, USA.

ABSTRACT
The detection and characterization of antigen-specific T cell populations is critical for understanding the development and physiology of the immune system and its responses in health and disease. We have developed and tested a method that uses arrays of peptide-MHC complexes for the rapid identification, isolation, activation, and characterization of multiple antigen-specific populations of T cells. CD4(+) or CD8(+) lymphocytes can be captured in accordance with their ligand specificity using an array of peptide-MHC complexes printed on a film-coated glass surface. We have characterized the specificity and sensitivity of a peptide-MHC array using labeled lymphocytes from T cell receptor transgenic mice. In addition, we were able to use the array to detect a rare population of antigen-specific T cells following vaccination of a normal mouse. This approach should be useful for epitope discovery, as well as for characterization and analysis of multiple epitope-specific T cell populations during immune responses associated with viral and bacterial infection, cancer, autoimmunity, and vaccination.

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Related in: MedlinePlus

A Cellular Microarray(A) A schematic diagram illustrating T cell binding to its cognate peptide–MHC spot via peptide–MHC–TCR interactions (not drawn to scale).(B) Cells immobilized on the cellular microarray are immediately visible to inspection. When cell coverage exceeds approximately 20% confluency, the cluster becomes visible to the naked eye. Shown are cell clusters bound to several triplicate spots of either peptide–MHC complexes or antibodies. The image was taken using a digital camera. The arrows and legends indicate the locations and identities of each of the printed triplicates. Spot diameter and interspot distance are about 400 μm and 700 μm, respectively. Scale bar is 1.4 mm in length.
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pbio.0000065-g001: A Cellular Microarray(A) A schematic diagram illustrating T cell binding to its cognate peptide–MHC spot via peptide–MHC–TCR interactions (not drawn to scale).(B) Cells immobilized on the cellular microarray are immediately visible to inspection. When cell coverage exceeds approximately 20% confluency, the cluster becomes visible to the naked eye. Shown are cell clusters bound to several triplicate spots of either peptide–MHC complexes or antibodies. The image was taken using a digital camera. The arrows and legends indicate the locations and identities of each of the printed triplicates. Spot diameter and interspot distance are about 400 μm and 700 μm, respectively. Scale bar is 1.4 mm in length.

Mentions: MHC tetramers and antibodies were arrayed using a piezo-electric noncontact arrayer, which minimizes film disruption and allows control over spot size and the quantity of material deposited, by enabling multiple nanoliter-scale drops to be deposited onto each spot. The resulting spot size with ten 0.45 nl drops (dispensed at a concentration of 1μg/μl) was approximately 400 μm in diameter. Each spot of this size could accommodate approximately 1,600 closely packed T cells. Suspended cells were subsequently layered onto the array and were allowed to interact with the predispensed tetramers and antibodies (Figure 1A). Following a short period of incubation, the slide was washed to remove unbound cells and visualized by direct inspection, differential interference contrast (DIC), or fluorescence microscopy. Inspection by eye revealed tight, nearly confluent cell clusters on both the antibody and the MHC tetramer spots (Figure 1B).


Detection and characterization of cellular immune responses using peptide-MHC microarrays.

Soen Y, Chen DS, Kraft DL, Davis MM, Brown PO - PLoS Biol. (2003)

A Cellular Microarray(A) A schematic diagram illustrating T cell binding to its cognate peptide–MHC spot via peptide–MHC–TCR interactions (not drawn to scale).(B) Cells immobilized on the cellular microarray are immediately visible to inspection. When cell coverage exceeds approximately 20% confluency, the cluster becomes visible to the naked eye. Shown are cell clusters bound to several triplicate spots of either peptide–MHC complexes or antibodies. The image was taken using a digital camera. The arrows and legends indicate the locations and identities of each of the printed triplicates. Spot diameter and interspot distance are about 400 μm and 700 μm, respectively. Scale bar is 1.4 mm in length.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC300678&req=5

pbio.0000065-g001: A Cellular Microarray(A) A schematic diagram illustrating T cell binding to its cognate peptide–MHC spot via peptide–MHC–TCR interactions (not drawn to scale).(B) Cells immobilized on the cellular microarray are immediately visible to inspection. When cell coverage exceeds approximately 20% confluency, the cluster becomes visible to the naked eye. Shown are cell clusters bound to several triplicate spots of either peptide–MHC complexes or antibodies. The image was taken using a digital camera. The arrows and legends indicate the locations and identities of each of the printed triplicates. Spot diameter and interspot distance are about 400 μm and 700 μm, respectively. Scale bar is 1.4 mm in length.
Mentions: MHC tetramers and antibodies were arrayed using a piezo-electric noncontact arrayer, which minimizes film disruption and allows control over spot size and the quantity of material deposited, by enabling multiple nanoliter-scale drops to be deposited onto each spot. The resulting spot size with ten 0.45 nl drops (dispensed at a concentration of 1μg/μl) was approximately 400 μm in diameter. Each spot of this size could accommodate approximately 1,600 closely packed T cells. Suspended cells were subsequently layered onto the array and were allowed to interact with the predispensed tetramers and antibodies (Figure 1A). Following a short period of incubation, the slide was washed to remove unbound cells and visualized by direct inspection, differential interference contrast (DIC), or fluorescence microscopy. Inspection by eye revealed tight, nearly confluent cell clusters on both the antibody and the MHC tetramer spots (Figure 1B).

Bottom Line: The detection and characterization of antigen-specific T cell populations is critical for understanding the development and physiology of the immune system and its responses in health and disease.We have developed and tested a method that uses arrays of peptide-MHC complexes for the rapid identification, isolation, activation, and characterization of multiple antigen-specific populations of T cells.In addition, we were able to use the array to detect a rare population of antigen-specific T cells following vaccination of a normal mouse.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Stanford University, Stanford, California, USA.

ABSTRACT
The detection and characterization of antigen-specific T cell populations is critical for understanding the development and physiology of the immune system and its responses in health and disease. We have developed and tested a method that uses arrays of peptide-MHC complexes for the rapid identification, isolation, activation, and characterization of multiple antigen-specific populations of T cells. CD4(+) or CD8(+) lymphocytes can be captured in accordance with their ligand specificity using an array of peptide-MHC complexes printed on a film-coated glass surface. We have characterized the specificity and sensitivity of a peptide-MHC array using labeled lymphocytes from T cell receptor transgenic mice. In addition, we were able to use the array to detect a rare population of antigen-specific T cells following vaccination of a normal mouse. This approach should be useful for epitope discovery, as well as for characterization and analysis of multiple epitope-specific T cell populations during immune responses associated with viral and bacterial infection, cancer, autoimmunity, and vaccination.

Show MeSH
Related in: MedlinePlus