Optical coding of mammalian cells using semiconductor quantum dots.

Cell-based assays are widely used to screen compounds and study complex phenotypes. Few methods exist, however, for multiplexing cellular assays or labeling individual cells in a mixed cell population. We developed a generic encoding method for cells that is based on peptide-mediated delivery of quantum dots (QDs) into live cells. The QDs are nontoxic and photostable and can be imaged using conventional fluorescence microscopy or flow cytometry systems. We created unique fluorescent codes for a variety of mammalian cell types and show that our encoding method has the potential to create > 100 codes. We demonstrate that QD cell codes are compatible with most types of compound screening assays including immunostaining, competition binding, reporter gene, receptor internalization, and intracellular calcium release. A multiplexed calcium assay for G-protein-coupled receptors using QDs is demonstrated. The ability to spectrally encode individual cells with unique fluorescent barcodes should open new opportunities in multiplexed assay development and greatly facilitate the study of cell/cell interactions and other complex phenotypes in mixed cell populations.

[1]  S. P. Fodor,et al.  Light-directed, spatially addressable parallel chemical synthesis. , 1991, Science.

[2]  S. Schreiber,et al.  Printing proteins as microarrays for high-throughput function determination. , 2000, Science.

[3]  D. Balding,et al.  HLA Sequence Polymorphism and the Origin of Humans , 2006 .

[4]  D. S. Gill,et al.  Optical multibead arrays for simple and complex odor discrimination. , 2001, Analytical chemistry.

[5]  J R Kettman,et al.  Multiplexed analysis of human cytokines by use of the FlowMetrix system. , 1998, Clinical chemistry.

[6]  M. Mrksich,et al.  Using electroactive substrates to pattern the attachment of two different cell populations , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[7]  R. G. Freeman,et al.  Submicrometer metallic barcodes. , 2001, Science.

[8]  Milan Mrksich,et al.  Micropatterned Surfaces for Control of Cell Shape, Position, and Function , 1998, Biotechnology progress.

[9]  S. Pathak,et al.  Hydroxylated quantum dots as luminescent probes for in situ hybridization. , 2001, Journal of the American Chemical Society.

[10]  D E Ingber,et al.  Using microcontact printing to pattern the attachment of mammalian cells to self-assembled monolayers of alkanethiolates on transparent films of gold and silver. , 1997, Experimental cell research.

[11]  J. Matthew Mauro,et al.  Long-term multiple color imaging of live cells using quantum dot bioconjugates , 2003, Nature Biotechnology.

[12]  D. Sabatini,et al.  Microarrays of cells expressing defined cDNAs , 2001, Nature.

[13]  M. Weiner,et al.  A microsphere-based assay for multiplexed single nucleotide polymorphism analysis using single base chain extension. , 2000, Genome research.

[14]  James McBride,et al.  Targeting cell surface receptors with ligand-conjugated nanocrystals. , 2002, Journal of the American Chemical Society.

[15]  S. Nie,et al.  Quantum-dot-tagged microbeads for multiplexed optical coding of biomolecules , 2001, Nature Biotechnology.

[16]  M. Bruchez,et al.  Immunofluorescent labeling of cancer marker Her2 and other cellular targets with semiconductor quantum dots , 2003, Nature Biotechnology.

[17]  Vincent Noireaux,et al.  In Vivo Imaging of Quantum Dots Encapsulated in Phospholipid Micelles , 2002, Science.

[18]  D E Ingber,et al.  Controlling cell attachment on contoured surfaces with self-assembled monolayers of alkanethiolates on gold. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[19]  H. Lehrach,et al.  Protein microarrays for gene expression and antibody screening. , 1999, Analytical biochemistry.

[20]  A. Alivisatos.  Semiconductor Clusters, Nanocrystals, and Quantum Dots , 1996, Science.

[21]  M. Bawendi,et al.  (CdSe)ZnS Core-Shell Quantum Dots - Synthesis and Characterization of a Size Series of Highly Luminescent Nanocrystallites , 1997 .

[22]  T. Mosmann.  Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. , 1983, Journal of immunological methods.

[23]  A Paul Alivisatos,et al.  Sorting fluorescent nanocrystals with DNA. , 2002, Journal of the American Chemical Society.

[24]  J. Kauer,et al.  Convergent, self-encoded bead sensor arrays in the design of an artificial nose. , 1999, Analytical chemistry.

[25]  I. Tomlinson,et al.  Antibody arrays for high-throughput screening of antibody–antigen interactions , 2000, Nature Biotechnology.

[26]  S. Nie,et al.  Quantum dot bioconjugates for ultrasensitive nonisotopic detection. , 1998, Science.

[27]  M. Morris,et al.  A peptide carrier for the delivery of biologically active proteins into mammalian cells , 2001, Nature Biotechnology.

[28]  P. Guyot-Sionnest,et al.  Synthesis and Characterization of Strongly Luminescing ZnS-Capped CdSe Nanocrystals , 1996 .

[29]  Hedi Mattoussi,et al.  Avidin: a natural bridge for quantum dot-antibody conjugates. , 2002, Journal of the American Chemical Society.

[30]  D. Stites,et al.  Development of a microsphere-based fluorescent immunoassay and its comparison to an enzyme immunoassay for the detection of antibodies to three antigen preparations from Candida albicans. , 1989, Journal of immunological methods.

[31]  Christine M. Micheel,et al.  Cell Motility and Metastatic Potential Studies Based on Quantum Dot Imaging of Phagokinetic Tracks , 2002 .

[32]  D R Walt,et al.  Application of high-density optical microwell arrays in a live-cell biosensing system. , 2000, Analytical biochemistry.