title: Single-cell microarray for analyzing cellular response.
abstract: Detection of cellular response by measuring intracellular calcium, (Ca2+)i with Ca2+-dependent fluorescent dye are standard approaches to detect ligand-stimulated cells and to study signaling through ligand/receptor interaction. We describe a single-cell microarray system to analyze cellular response of individual cells such as lymphocytes using microchamber array chips. The single-cell microarray chip is made from polystyrene with over 30,000 microchambers, which can accommodate only single cells. Lymphocytes derived from mouse spleen or human blood were spread on the microarray, and over 80% of the microchambers achieved single-cell status. Stimulation of B-cells through antigen receptors on the microarray allowed us to detect activated B-cells by comparing the states of single B-cells before and after stimulation with antigen, which is disabled for flow cytometry. In addition, this novel method demonstrated retrieval of positive single B-cells from microchambers by a micromanipulator and achieved antibody DNA analysis. The system is suitable for high-throughput analysis of intracellular Ca2+ response at the single-cell level and is applicable to screen antigen-specific lymphocytes for making specific monoclonal antibody.
authors.affiliation: Toyama New Industry Organization, 529, Takada, Toyama 930-0866, and School of Materials Science, Japan Advanced Institute of Science and Technology, 1-1, Asahidai, Nomi City, Ishikawa 923-1292, Japan.
authors.forename: Shohei
authors.initials: S
authors.lastname: Yamamura
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kishi
authors.forename: Yoshiharu
authors.initials: Y
authors.lastname: Tokimitsu
authors.forename: Sachiko
authors.initials: S
authors.lastname: Kondo
authors.forename: Ritsu
authors.initials: R
authors.lastname: Honda
authors.forename: Sathuluri Ramachandra
authors.initials: SR
authors.lastname: Rao
authors.forename: Masahiro
authors.initials: M
authors.lastname: Omori
authors.forename: Eiichi
authors.initials: E
authors.lastname: Tamiya
authors.forename: Atsushi
authors.initials: A
authors.lastname: Muraguchi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology; D001483:Base Sequence; D002118:Calcium / Q000032:analysis; D020013:Calcium Signaling; D006801:Humans; D007075:Immunoglobulin M / Q000737:chemistry / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000737:chemistry; D008214:Lymphocytes / Q000378:metabolism*; D051379:Mice; D046228:Microarray Analysis / Q000295:instrumentation* / Q000379:methods*; D008969:Molecular Sequence Data
chemical_list: D007075:Immunoglobulin M; D007135:Immunoglobulin Variable Region; D002118:Calcium