Single-cell microarray for analyzing cellular response.
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.
Yamamura
Shohei
S
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.
Kishi
Hiroyuki
H
Tokimitsu
Yoshiharu
Y
Kondo
Sachiko
S
Honda
Ritsu
R
Rao
Sathuluri Ramachandra
SR
Omori
Masahiro
M
Tamiya
Eiichi
E
Muraguchi
Atsushi
A
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
D007075:Immunoglobulin M; D007135:Immunoglobulin Variable Region; D002118:Calcium