Single-cell immunosensors for protein detection.
A single-cell detector is described that combines the natural signal amplification of whole-cell biosensors with the flexibility and specificity of immunological recognition. An immune cell that expresses receptors for the constant region of immunoglobulin G (IgG) is loaded with a Ca(2+)-indicating dye and with antibodies directed against the protein of interest. Introduction of a multivalent protein antigen causes cross-linking of the receptors, which results in a detectable increase in the concentration of cytosolic Ca(2+). Some immune cell lines respond to stimulation with oscillations in their cytosolic Ca(2+) levels that complicate their use as detectors. The human monocytic cell line U-937, when treated with the cytokine interferon-gamma, produces a large, short-lived Ca(2+) signal in response to cross-linking of its high-affinity IgG receptors. U-937 was therefore chosen for development as an immunity-based detector. Human and rabbit antibodies are found to effectively stimulate the cell, causing a prompt and transient response. The cell is able to respond to repeated stimulation, though the response diminishes during rapid stimulation. Ovalbumin can be detected in micromolar concentrations. Possible fundamental constraints on the size of a detectable analyte are discussed.
Whelan
Rebecca J
RJ
Department of Chemistry, Stanford University, Stanford, CA 94305-5080, USA.
Zare
Richard N
RN
D015374:Biosensing Techniques / Q000379:methods*; D002118:Calcium / Q000032:analysis*; D018929:Cell Culture Techniques / Q000379:methods*; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D010047:Ovalbumin / Q000032:analysis* / Q000276:immunology; D011506:Proteins / Q000032:analysis* / Q000276:immunology; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000379:methods*; D020298:U937 Cells / Q000502:physiology*
D011506:Proteins; D010047:Ovalbumin; D002118:Calcium