Immuno-Hybridization Chain Reaction for Enhancing Detection of Individual Cytokine-Secreting Human Peripheral Mononuclear Cells

Abstract

We present here a new method to enhance the detection of secreted cytokines and chemokines from single human mononuclear cells. The technique uses a hybridization chain reaction (HCR) to amplify signals resulting from sandwich immunoassays. This immuno-HCR employs oligonucleotide-based initiators covalently linked to antibodies to propagate a chain reaction of hybridization events involving a pair of complementary hairpin oligomers bearing fluorescent labels. Integrating this strategy for signal amplification with microengraving—a soft lithographic method for printing arrays of secreted proteins from thousands of single cells—improves both the limits of detection and sensitivity for cytokines and chemokines captured from individual cells by an average of 200-fold relative to methods for direct detection by fluoresence. This approach should enhance the utility of microengraving for defining the immunological signatures of diseases and responses to interventional therapies based on multiplexed single-cell analysis.

Enhanced electrochemical recognition of double-stranded DNA by using hybridization chain reaction and positively charged gold nanoparticles

“…Unlike other amplification approaches, HCR owns significant advantages including enzyme free, low background and PCR-like sensitivity (Ge et al, 2014;Huang et al, 2011). These characteristics make HCR be a promising candidate for the detection of DNA Chen et al, 2012;Niu et al, 2010;Bi et al, 2013;Li et al, 2014;Qian et al, 2015;Ren et al, 2015), protein (Song et al, 2014;Zhou et al, 2012;Dai et al, 2012;Zhang et al, 2012), metal ions Yuan et al, 2011;, cell (Choi et al, 2011) and small molecules by using electrochemical, fluorescent, colorimetric methods and so on.…”

Mass effect of redox reactions: A novel mode for surface plasmon resonance-based bioanalysis

“…2A. The high molecular weights, reflected as the bright band in Lanes 1 and 2 with a distinctive tailing, is indicative of the successful HCR with various lengths of chain-ladder dsDNA, which would overall dedicate to enormous signal amplification (Choi et al, 2011). In the meantime, the progress of HCR was kept tracked of on the SPR platform (Fig.…”

Target-catalyzed hairpin assembly and intramolecular/intermolecular co-reaction for signal amplified electrochemiluminescent detection of microRNA

“…Prior to use, the DNA hairpin oligonucleotides were denatured for 2 min at 95 ºC and slowly cooled to room temperature to form stem-loop structures (Choi et al, 2011).…”