Single-molecule detection of protein efflux from microorganisms using fluorescent single-walled carbon nanotube sensor arrays.
Markita Patricia LandryHiroki AndoAllen Y ChenJicong CaoVishal Isaac KottadielLinda ChioDarwin YangJuyao DongTimothy K LuMichael S StranoPublished in: Nature nanotechnology (2017)
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.
Keyphrases
- label free
- escherichia coli
- single cell
- amino acid
- protein protein
- single molecule
- binding protein
- stem cells
- high throughput
- high resolution
- cell death
- small molecule
- living cells
- antiretroviral therapy
- high density
- signaling pathway
- pseudomonas aeruginosa
- oxidative stress
- induced apoptosis
- bone marrow
- hiv positive
- real time pcr
- walled carbon nanotubes
- men who have sex with men
- cell proliferation
- hiv aids
- cystic fibrosis
- fluorescent probe
- hiv testing
- atomic force microscopy
- cell therapy