Single-molecule detection of protein efflux from microorganisms using fluorescent single-walled carbon nanotube sensor arrays.
Single-molecule detection of protein efflux from microorganisms using fluorescent single-walled carbon nanotube sensor arrays.
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DOI：
10.1038/nnano.2016.284
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发表时间：
2017-05
影响因子：
38.3
通讯作者：
Strano MS
中科院分区：
文献类型：
--
作者：
Landry MP;Ando H;Chen AY;Cao J;Kottadiel VI;Chio L;Yang D;Dong J;Lu TK;Strano MS
纳米传感器阵列的一个明显优势是，通过接近分析物的放置，它们能够在溶液中实现超低的检测限。在这里，我们展示了在微流控室中固定的大肠杆菌（细菌）和毕赤酵母（酵母）的单个蛋白质的无标记检测，实时测量单个生物的蛋白质外溢。该阵列采用非共价偶联的方式将适配体锚定多核苷酸序列与近红外发射单壁碳纳米管结合，并使用可变化学间隔剂优化传感器响应。未标记的RAP1 GTPase和HIV整合酶蛋白通过大的近红外荧光开启反应从各种细胞系中选择性地检测到。我们发现大肠杆菌诱导、蛋白质合成和蛋白质输出的过程是高度随机的，产生蛋白质分泌的可变性，在饥饿条件下进行分裂的大肠杆菌细胞分泌的蛋白质产物比非分裂的大肠杆菌细胞少66%。我们进一步展示了对大肠杆菌T7噬菌体感染产生的独特蛋白质产物的检测，表明纳米传感器阵列可以实时、单细胞地分析来自各种细胞类型的广泛蛋白质产物。
A distinct advantage of nanosensor arrays is their ability to achieve ultra-low 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. Unlabeled RAP1 GTPase and HIV integrase proteins were selectively detected from various cell lines, via large near-infrared fluorescent turn-on responses. We show 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.
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DOI：
10.1021/jp511448e
发表时间：
2015-05-07
期刊：
The journal of physical chemistry. C, Nanomaterials and interfaces
影响因子：
--
作者：
Landry MP;Vuković L;Kruss S;Bisker G;Landry AM;Islam S;Jain R;Schulten K;Strano MS
通讯作者：
Strano MS
影响因子：
10.8
作者：
Ament, Irene;Prasad, Janak;Soennichsen, Carsten
通讯作者：
Soennichsen, Carsten
影响因子：
30.8
作者：
Lahav, G;Rosenfeld, N;Alon, U
通讯作者：
Alon, U
影响因子：
17.1
作者：
Hammock, Mallory L.;Knopfmacher, Oren;Bao, Zhenan
通讯作者：
Bao, Zhenan
影响因子：
8.4
作者：
Keasling, Jay D.
通讯作者：
Keasling, Jay D.