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Published July 22, 2009 | Supplemental Material + Accepted Version
Journal Article Open

Modular Nucleic Acid Assembled p/MHC Microarrays for Multiplexed Sorting of Antigen-Specific T Cells


The human immune system consists of a large number of T cells capable of recognizing and responding to antigens derived from various sources. The development of peptide-major histocompatibility (p/MHC) tetrameric complexes has enabled the direct detection of these antigen-specific T cells. With the goal of increasing throughput and multiplexing of T cell detection, protein microarrays spotted with defined p/MHC complexes have been reported, but studies have been limited due to the inherent instability and reproducibility of arrays produced via conventional spotted methods. Herein, we report on a platform for the detection of antigen-specific T cells on glass substrates that offers significant advantages over existing surface-bound schemes. In this approach, called "Nucleic Acid Cell Sorting (NACS)", single-stranded DNA oligomers conjugated site-specifically to p/MHC tetramers are employed to immobilize p/MHC tetramers via hybridization to a complementary-printed substrate. Fully assembled p/MHC arrays are used to detect and enumerate T cells captured from cellular suspensions, including primary human T cells collected from cancer patients. NACS arrays outperform conventional spotted arrays assessed in key criteria such as repeatability and homogeneity. The versatility of employing DNA sequences for cell sorting is exploited to enable the programmed, selective release of target populations of immobilized T cells with restriction endonucleases for downstream analysis. Because of the performance, facile and modular assembly of p/MHC tetramer arrays, NACS holds promise as a versatile platform for multiplexed T cell detection.

Additional Information

© 2009 American Chemical Society. Received February 3, 2009; Publication Date (Web): June 24, 2009. The authors thank Takeshi Sano at Harvard Medical School for providing the pET-3a-SAC plasmid, John Altman at Emory University for providing an initial aliquot of SAC for trial experiments. We also thank Bruz Marzolf at the Institute for Systems Biology for printing the DNA microarrays, Mireille Riedinger at UCLA for assistance with SAC expression, Lilah Morris M.D. at UCLA for assistance with primary murine cell purifications. K.H. is supported by a Samsung Fellowship. O.N.W. is an Investigator of the Howard Hughes Medical Institute. This work was primarily funded by the National Cancer Institute, Grant No. 5U54 CA119347 (J.R.H., P.I.). Flow cytometry was performed in the UCLA Jonsson Comprehensive Cancer Center and Center for AIDS Research Flow Cytometry Core Facility, which is supported by the National Institutes of Health Awards CA-16042 and AI_28697, by the Jonsson Cancer Center, the UCLA AIDS Institute, and the UCLA School of Medicine.

Attached Files

Accepted Version - nihms127833.pdf

Supplemental Material - ja9006707_si_001.pdf


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