Biosensor for direct cell detection, quantification and analysis.

Microarrays are promising tools for cell isolation and detection. However, they have yet to be widely applied in biology. This stems from a lack of demonstration of their sensitivity and compatibility with complex biological samples, and a lack of proof that their use does not induce aberrant cellular effects. Herein, we characterized and optimized a recently developed technology associating antibody microarrays with surface plasmon resonance imaging (SPRi). Using a murine macrophage cell line we demonstrate the binding specificity of our antibody-microarrays and the correlation between SPRi signals and both the number of bound cells, and the level of expression of cell surface markers. Confocal microscopy reveals that cell binding to the chip through antibody-antigen interactions underwent morphological changes reflecting the density of the relevant cell surface marker without affecting cell viability as shown by fluorescent microscopy. The detection threshold of the microarray-SPRi system is lowered 10-fold by applying a polyethylene oxide film to the gold surface of the chip. This increased sensitivity allows the detection of cells representing as little as 0.5% of a mixed population. The potential of this method is illustrated by two applications: characterization of ligand-cell receptor interactions, allowing determination of receptor specificity, and analysis of peripheral blood mononuclear cells, demonstrating the suitability of this tool for the analysis of complex biological samples.

[1]  L. Herzenberg,et al.  Antigen-specific identification and cloning of hybridomas with a fluorescence-activated cell sorter. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[2]  Mark M Davis,et al.  Detection and Characterizationof Cellular Immune Responses Using Peptide–MHC Microarrays , 2003, PLoS biology.

[3]  R. Purcell,et al.  CD8+ T Cells Mediate Viral Clearance and Disease Pathogenesis during Acute Hepatitis B Virus Infection , 2003, Journal of Virology.

[4]  H. Grey,et al.  Receptors for IgG: subclass specificity of receptors on different mouse cell types and the definition of two distinct receptors on a macrophage cell line , 1977, The Journal of experimental medicine.

[5]  Wolfgang Knoll,et al.  Surface–plasmon microscopy , 1988, Nature.

[6]  Mark M Davis,et al.  Marked Differences in Human Melanoma Antigen-Specific T Cell Responsiveness after Vaccination Using a Functional Microarray , 2005, PLoS medicine.

[7]  B. Liedberg,et al.  Surface plasmon resonance for gas detection and biosensing , 1983 .

[8]  Günter Gauglitz,et al.  Surface plasmon resonance sensors: review , 1999 .

[9]  P. Marche,et al.  Simultaneous evaluation of lymphocyte subpopulations in the liver and in peripheral blood mononuclear cells of HCV‐infected patients: relationship with histological lesions , 2002, Clinical and experimental immunology.

[10]  T. Livache,et al.  Real-time detection of lymphocytes binding on an antibody chip using SPR imaging. , 2007, Lab on a chip.

[11]  C. Borrebaeck,et al.  Antibody microarrays: current status and key technological advances. , 2006, Omics : a journal of integrative biology.

[12]  T. Livache,et al.  Peptide-protein microarrays and surface plasmon resonance detection: biosensors for versatile biomolecular interaction analysis. , 2010, Biosensors & bioelectronics.

[13]  H. Young,et al.  Immortalized Myeloid Suppressor Cells Trigger Apoptosis in Antigen-Activated T Lymphocytes1 , 2000, The Journal of Immunology.

[14]  C. Verdier,et al.  Measuring cell viscoelastic properties using a force-spectrometer: influence of protein-cytoplasm interactions. , 2005, Biorheology.

[15]  F. Rossi,et al.  Fabrication and characterization of protein arrays for stem cell patterning , 2009 .

[16]  Michael R Shortreed,et al.  Specific capture of mammalian cells by cell surface receptor binding to ligand immobilized on gold thin films. , 2006, Journal of proteome research.

[17]  F. Rossi,et al.  Fouling and non-fouling surfaces produced by plasma polymerization of ethylene oxide monomer. , 2006, Acta biomaterialia.

[18]  Dan Davidov,et al.  Infrared surface plasmon resonance: a novel tool for real time sensing of variations in living cells. , 2006, Biophysical journal.

[19]  M. Raff,et al.  Redistribution and pinocytosis of lymphocyte surface immunoglobulin molecules induced by anti-immunoglobulin antibody. , 1971, Nature: New biology.

[20]  GEOFFREY TAYLOR,et al.  Aeronautics before 1919 , 1971, Nature.

[21]  P. Marche,et al.  A new role for complement C3: regulation of antigen processing through an inhibitory activity. , 2008, Molecular immunology.

[22]  R. Dörries.  The role of T-cell-mediated mechanisms in virus infections of the nervous system. , 2001, Current topics in microbiology and immunology.

[23]  Thierry Livache,et al.  Clinically related protein-peptide interactions monitored in real time on novel peptide chips by surface plasmon resonance imaging. , 2006, Clinical chemistry.

[24]  Lucel Sirghi,et al.  Micro-stamped surfaces for the patterned growth of neural stem cells. , 2008, Biomaterials.

[25]  P. Klenerman,et al.  T cell sensitivity and the outcome of viral infection , 2010, Clinical and experimental immunology.

[26]  Hidenori Suzuki,et al.  The SPR signal in living cells reflects changes other than the area of adhesion and the formation of cell constructions. , 2007, Biosensors & bioelectronics.

[27]  R. Guy,et al.  A High-Throughput Ligand Competition Binding Assay for the Androgen Receptor and Other Nuclear Receptors , 2009, Journal of biomolecular screening.

[28]  M. Greaves,et al.  Detection of refractive index changes in individual living cells by means of surface plasmon resonance imaging. , 2010, Biosensors & bioelectronics.

[29]  Thierry Livache,et al.  A polypyrrole protein microarray for antibody-antigen interaction studies using a label-free detection process. , 2005, Analytical biochemistry.