Single-molecule detection of protein efflux from microorganisms using fluorescent single-walled carbon nanotube sensor arrays

Abstract

A distinct advantage of nanosensor arrays is their ability to achieve ultralow detection limits in solution by proximity placement to an analyte. Here, we demonstrate label-free detection of individual proteins from Escherichia coli (bacteria) and Pichia pastoris (yeast) immobilized in a microfluidic chamber, measuring protein efflux from single organisms in real time. The array is fabricated using non-covalent conjugation of an aptamer-anchor polynucleotide sequence to near-infrared emissive single-walled carbon nanotubes, using a variable chemical spacer shown to optimize sensor response. Unlabelled RAP1 GTPase and HIV integrase proteins were selectively detected from various cell lines, via large near-infrared fluorescent turn-on responses. We show that the process of E. coli induction, protein synthesis and protein export is highly stochastic, yielding variability in protein secretion, with E. coli cells undergoing division under starved conditions producing 66% fewer secreted protein products than their non-dividing counterparts. We further demonstrate the detection of a unique protein product resulting from T7 bacteriophage infection of E. coli, illustrating that nanosensor arrays can enable real-time, single-cell analysis of a broad range of protein products from various cell types.

DNA–Carbon Nanotube Complexation Affinity and Photoluminescence Modulation Are Independent

“…10 The ability to integrate molecular recognition into DNA-nanotube hybrids has been achieved via base-pair hybridization, 11 antibody-coupling, 12 and oligonucleotide aptamers. 13 …”

Recent Advances and Perspectives in Microfluidics‐Based Single‐Cell Biosensing Techniques

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Chemometric Approaches for Developing Infrared Nanosensors to Image Anthracyclines

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