The confined geometry of nanopores enables a wealth of chemistry and analysis to be conducted at the single-molecule scale. Yi-Lun Ying, Aleksandar P. Ivanov and Vincent Tabard-Cossa report on recent developments discussed at the 2020 Nanopore Electrochemistry Meeting.納米孔的受限幾何形狀使得能夠在單分子規(guī)模上進(jìn)行大量的化學(xué)和分析。 應(yīng)義倫，Aleksandar P.Ivanov和Vincent Tabard-Cossa報(bào)告了在2020年納米孔電化學(xué)會(huì)議上討論的最新進(jìn)展窒悔。

One of the most significant challenges in modern analytical sciences is the ever-growing need to characterize single-molecules and molecular assemblies with high spatial and temporal resolution. Nanopores are nanoscale-sized channels that address this challenge by providing a confined space for detecting single entities such as small molecules, nucleic acid polymers, proteins, viruses and nanoparticles using an electrical signal1,2,3. When a voltage is applied across the nanopore, its confined geometry forms a 3D sensing interface that is probed continuously by an ionic current. Molecules that are temporarily confined or transported through the nanopore can be identified by their current modulation. Various methods could be used to controllably construct nanopore structures in a range of soft and solid-state materials. These may include the self-assembly of proteins in lipid bilayers or block copolymer membranes, the drilling or dielectric breakdown of solid-state membranes, or even by simple pulling of glass capillaries down to nanoscale dimensions3,4.現(xiàn)代分析科學(xué)中最重大的挑戰(zhàn)之一是對(duì)以高時(shí)空分辨率表征單分子和分子組裝體的需求不斷增長(zhǎng)俊性。納米孔是納米級(jí)通道堤框，通過(guò)提供一個(gè)狹窄的空間來(lái)利用電信號(hào)1,2,3檢測(cè)單個(gè)實(shí)體朋蔫，例如小分子找都，核酸聚合物萝究，蛋白質(zhì)免都，病毒和納米顆粒，從而解決了這一難題帆竹。當(dāng)在納米孔上施加電壓時(shí)绕娘，其有限的幾何形狀會(huì)形成3D感應(yīng)界面，該界面會(huì)被離子電流連續(xù)探測(cè)栽连∠樟欤可以通過(guò)電流調(diào)制來(lái)識(shí)別暫時(shí)限制或轉(zhuǎn)運(yùn)通過(guò)納米孔的分子∶虢簦可以使用各種方法來(lái)可控地構(gòu)建一系列軟和固態(tài)材料中的納米孔結(jié)構(gòu)绢陌。這些可能包括蛋白質(zhì)在脂質(zhì)雙層或嵌段共聚物膜中的自組裝，固態(tài)膜的鉆孔或介電擊穿熔恢，甚至可以通過(guò)簡(jiǎn)單地將玻璃毛細(xì)管拉至納米尺寸3,4來(lái)完成脐湾。

By confining single molecules, nanopore sensors could access information on the chemical and physical properties of molecular entities, which is remarkably wealthier than only looking at ensemble-average measurements. The approach has already proven its prowess for long-read nucleic acid sequencing and is making its first strides towards the formidable goal of achieving protein sequencing. Beyond sequencing, it offers an elegant solution to a diverse array of contemporary challenges, for example, by providing novel insights into outstanding ionic and molecular transport problems, controllable chemical synthesis, precision diagnostics and advances in green energy conversion.通過(guò)限制單個(gè)分子，納米孔傳感器可以訪問(wèn)有關(guān)分子實(shí)體的化學(xué)和物理性質(zhì)的信息叙淌，這比僅查看集合平均測(cè)量值要豐富得多秤掌。該方法已經(jīng)證明了其在長(zhǎng)時(shí)間閱讀核酸測(cè)序中的能力，并且正在朝著實(shí)現(xiàn)蛋白質(zhì)測(cè)序的強(qiáng)大目標(biāo)邁出第一步鹰霍。除測(cè)序之外机杜，它還為各種當(dāng)代挑戰(zhàn)提供了一個(gè)優(yōu)雅的解決方案，例如衅谷，通過(guò)提供對(duì)未解決的離子和分子傳輸問(wèn)題，可控化學(xué)合成似将，精確診斷和綠色能源轉(zhuǎn)換方面的新見(jiàn)解获黔。

The 2020 Nanopore Electrochemistry Meeting (9–14th October) provided an opportunity for scientists worldwide to share recent advances and exchange ideas on the growing number of potential applications that continuously drive the rapid development of nanopore analysis (Fig. 1). The virtual meeting was hosted and organized by Yi-Tao Long and Hong-Yuan Chen from Nanjing University, China and Mathias Winterhalter from Jacobs University, Germany. Over 800 participants from across the globe came together to attend over 60 presentations and discuss more than 1000 questions online. These discussions covered an extensive range of nanopore-related topics spanning across physical and life sciences, medicine and engineering. The speakers shared new insights on biological suprabiomolecular assemblies, bionics, functional (bio)materials, chemical reactions and unconventional transport in nano-confined environments. There was significant focus on emerging nanopore methods for controllable chemical synthesis, protein analysis, future precision diagnostics using molecular barcodes and carriers, biosensing and molecular information storage using DNA nanostructures, single-cell analysis, and for providing new insights into the dynamics of biological processes.2020年納米孔電化學(xué)會(huì)議（10月9日至14日）為全世界的科學(xué)家提供了一個(gè)分享最新進(jìn)展并就不斷推動(dòng)納米孔分析快速發(fā)展的潛在應(yīng)用交流想法的機(jī)會(huì)（圖1）蚀苛。虛擬會(huì)議由中國(guó)南京大學(xué)的龍逸濤和陳洪元以及德國(guó)雅各布斯大學(xué)的Mathias Winterhalter主持和組織。來(lái)自全球的800多名與會(huì)者聚集在一起玷氏，參加了60多個(gè)演講堵未，并在線討論了1000多個(gè)問(wèn)題。這些討論涵蓋了與納米孔相關(guān)的廣泛主題盏触，涉及物理和生命科學(xué)渗蟹，醫(yī)學(xué)和工程學(xué)。演講者分享了有關(guān)生物超分子組裝赞辩，仿生學(xué)雌芽，功能性（生物）材料，化學(xué)反應(yīng)和納米受限環(huán)境中非常規(guī)運(yùn)輸?shù)男乱?jiàn)解辨嗽。人們非常關(guān)注新興的納米孔方法世落，這些方法可用于可控制的化學(xué)合成，蛋白質(zhì)分析糟需，使用分子條形碼和載體的未來(lái)精確診斷屉佳，使用DNA納米結(jié)構(gòu)的生物傳感和分子信息存儲(chǔ)，單細(xì)胞分析以及提供對(duì)生物過(guò)程動(dòng)力學(xué)的新見(jiàn)解洲押。?

A new emerging domain in measurement sciences, nanopore electrochemistry, uses a nanopore to characterize single entities with spatial resolution down to sub-nanometre scale and temporal resolution reaching sub-microseconds. The electrochemical confinement of protons, ions, charges, analytes, reactions, interactions and fluidics inside nanopore channels offers a powerful toolkit for the analytical sciences and provides new insights into outstanding scientific questions in protein science, controllable chemical synthesis, and precision diagnostics.

測(cè)量科學(xué)中一個(gè)新興領(lǐng)域是納米孔電化學(xué)武花，它使用納米孔來(lái)表征單個(gè)實(shí)體，其空間分辨率低至亞納米級(jí)杈帐，時(shí)間分辨率達(dá)到亞微秒体箕。納米孔通道內(nèi)質(zhì)子，離子娘荡，電荷干旁，分析物，反應(yīng)炮沐，相互作用和流體的電化學(xué)限制為分析科學(xué)提供了強(qiáng)大的工具包争群，并提供了對(duì)蛋白質(zhì)科學(xué)，可控化學(xué)合成和精確診斷中杰出科學(xué)問(wèn)題的新見(jiàn)解大年。

One exciting trend was the growing versatility of biological nanopores. The hallmark of the protein nanopore is the atomically defined interior, which offers a designable bioreaction interface and excellent control of the local environment. Hagan Bayley’s team (University of Oxford, UK) shared their latest results using an α-hemolysin nanopore as a “nanoreactor” for single-molecule covalent chemistry. When an analyte molecule covalently attaches to the reactive site on the pore interior wall, the chemical reaction can be inferred by a change in ionic current. The intermediates in a reaction can be registered and their lifetimes recorded, yielding the rate constant for all steps. In an organic chemistry context, Scott L. Cockroft (University of Edinburgh, UK) showed that nanopores could resolve rapid click reaction equilibria. Henry S. White (University of Utah, USA) discussed acid–base chemistry in base-flipping dynamics. Learning from the quantum-confined superfluid (QSF) effect in biological ion channels, Lei Jiang’s group (Technical Institute of Physics and Chemistry, China) aimed to arrange single molecules in artificial nanopores to improve the efficiency of chemical reactions. Yunfei Chen (Southeast University, China) discussed dehydrated ion interactions in a highly confined nanopore solution. Chemistry under nanopore confinement thus permits many notable applications, including in situ site-specific modification of biopolymers, selective transmembrane signalling in synthetic cells and enantioselective catalysis. Nanopore confinement also enables the control of electrochemical phenomena such as permselective ion transport (when an ion is preferentially transported), gated transport and redox cycling. To this end, Paul W. Bohn (University of Notre Dame, USA) showed hierarchically organized nanopore electrode arrays to control the transport and reactivity of redox molecules.

一種令人興奮的趨勢(shì)是生物納米孔的多功能性不斷增長(zhǎng)换薄。蛋白質(zhì)納米孔的標(biāo)志是原子定義的內(nèi)部，提供了可設(shè)計(jì)的生物反應(yīng)界面和對(duì)本地環(huán)境的出色控制翔试。 Hagan Bayley的小組（英國(guó)牛津大學(xué)）分享了他們的最新結(jié)果轻要，他們使用α-溶血素納米孔作為單分子共價(jià)化學(xué)的“納米反應(yīng)器”。當(dāng)分析物分子共價(jià)附于孔內(nèi)壁上的反應(yīng)位點(diǎn)時(shí)垦缅，可以通過(guò)離子電流的變化來(lái)推斷化學(xué)反應(yīng)冲泥。可以記錄反應(yīng)中的中間體并記錄其壽命，從而獲得所有步驟的速率常數(shù)凡恍。在有機(jī)化學(xué)背景下志秃，Scott L. Cockroft（英國(guó)愛(ài)丁堡大學(xué)）表明，納米孔可以解決快速點(diǎn)擊反應(yīng)的平衡問(wèn)題嚼酝。亨利·懷特（Henry S. White）（美國(guó)猶他大學(xué)）在堿翻轉(zhuǎn)動(dòng)力學(xué)中討論了酸堿化學(xué)浮还。蔣磊的小組從生物離子通道中的量子限制超流體（QSF）效應(yīng)中學(xué)到了東西，旨在將單個(gè)分子排列在人造納米孔中闽巩，以提高化學(xué)反應(yīng)的效率钧舌。陳云飛（中國(guó)東南大學(xué)）討論了在高度受限的納米孔溶液中的脫水離子相互作用。因此涎跨，在納米孔限制下的化學(xué)方法具有許多顯著的應(yīng)用洼冻，包括生物聚合物的原位定點(diǎn)修飾，合成細(xì)胞中的選擇性跨膜信號(hào)傳導(dǎo)和對(duì)映選擇性催化六敬。納米孔的限制還能夠控制電化學(xué)現(xiàn)象碘赖，例如全選擇性離子遷移（優(yōu)先遷移離子時(shí)），門(mén)控遷移和氧化還原循環(huán)外构。為此普泡，Paul W. Bohn（美國(guó)圣母大學(xué)）展示了分層組織的納米孔電極陣列，以控制氧化還原分子的轉(zhuǎn)運(yùn)和反應(yīng)性审编。

Typical biological nanopores also provide some of the most advanced detectors for proteomic analysis at the single-molecule level. Juan Pelta (University of Paris-Saclay, France) described steps toward nanopore sequencing of proteins using a wild-type aerolysin nanopore, a powerful sensor for size discrimination of peptides. Together with Jan C. Behrends (University of Freiburg, Germany), they revealed that the aerolysin nanopore could identify single amino acid differences within a polyarginine carrier. By tuning the two sensing regions inside a mutant aerolysin nanopore, Yi-Tao Long’s group (Nanjing University, China) demonstrated enhanced sensitivity of nanopores for identifying peptide post-translation modifications and DNA lesions. Giovanni Maglia (University of Groningen, the Netherlands) introduced a sequencing concept based on assembling a nanopore protein sequencer with a proteasome that unfolds the protein structures. Biological nanopores are naturally suitable to study protein–protein interactions (PPI) and protein conformational states. Liviu Movileanu (Syracuse University, USA) showed a sensor platform based on a tether, a peptide adaptor and a protein receptor that could identify low- and high-affinity PPI in a complex mixture. Proteins are folded into 3D shapes of different sizes and solid-state nanopores have diameters that can be tuned easily to study a wide range of protein structures and biomolecular complexes. Moreover, solid-state nanopores allow current recordings in harsh conditions such as high voltage (force), high temperature, extreme pH and strong denaturants. Cees Dekker (TU Delft, the Netherlands) presented “NEOtrap”, a technique for long-term trapping (minutes to hours) of individual proteins inside a solid-state nanopore. A DNA-origami nanosphere is docked to a lipid-coated nanopore, forming a confined region and a highly charged environment where proteins can be trapped via electro-osmotic flow. Michael Mayer (Université de Fribourg, Switzerland) shared his vision of using polymer and lipid-coated solid-state nanopores for high-resolution characterization of single protein amyloid particles. Sébastien Balme (Université de Montpellier, France) described how conical track-etched nanopores, with their high aspect ratio, are well suited to characterize protein protofibril growth or degradation.典型的生物納米孔還為單分子水平的蛋白質(zhì)組學(xué)分析提供了一些最先進(jìn)的檢測(cè)器撼班。 Juan Pelta（法國(guó)巴黎薩克萊大學(xué)）描述了使用野生型氣溶素納米孔對(duì)蛋白質(zhì)進(jìn)行納米孔測(cè)序的步驟，這是一種用于區(qū)分多肽大小的強(qiáng)大傳感器垒酬。他們與德國(guó)弗萊堡大學(xué)的Jan C. Behrends一起發(fā)現(xiàn)砰嘁，溶血素納米孔可以識(shí)別聚精氨酸載體內(nèi)的單個(gè)氨基酸差異。通過(guò)調(diào)節(jié)突變的溶菌素納米孔內(nèi)部的兩個(gè)感應(yīng)區(qū)域勘究，Yi-Tao Long的小組（中國(guó)南京大學(xué)）證明了納米孔對(duì)識(shí)別肽段翻譯后修飾和DNA損傷的敏感性增強(qiáng)矮湘。 Giovanni Maglia（荷蘭格羅寧根大學(xué)）介紹了一種測(cè)序概念，該概念的基礎(chǔ)是將納米孔蛋白質(zhì)測(cè)序儀與可折疊蛋白質(zhì)結(jié)構(gòu)的蛋白酶體組裝在一起口糕。生物納米孔自然適合研究蛋白質(zhì)-蛋白質(zhì)相互作用（PPI）和蛋白質(zhì)構(gòu)象狀態(tài)缅阳。 Liviu Movileanu（美國(guó)雪城大學(xué)）展示了一種基于系鏈，肽接頭和蛋白質(zhì)受體的傳感器平臺(tái)景描，該平臺(tái)可識(shí)別復(fù)雜混合物中的低親和力和高親和力PPI十办。蛋白質(zhì)被折疊成不同大小的3D形狀，并且固態(tài)納米孔的直徑可以輕松調(diào)整超棺，以研究各種蛋白質(zhì)結(jié)構(gòu)和生物分子復(fù)合物向族。而且，固態(tài)納米孔可以在苛刻的條件下記錄電流棠绘，例如高壓（力）件相，高溫再扭，極端的pH值和強(qiáng)變性劑。 Cees Dekker（荷蘭TU Delft）提出了“ NEOtrap”技術(shù)适肠，該技術(shù)可將固態(tài)納米孔中的單個(gè)蛋白質(zhì)長(zhǎng)期捕獲（數(shù)分鐘至數(shù)小時(shí)）霍衫。 DNA折紙納米球停靠在脂質(zhì)包裹的納米孔上侯养，形成一個(gè)狹窄的區(qū)域和一個(gè)高度帶電的環(huán)境，可以通過(guò)電滲流捕獲蛋白質(zhì)澄干。 Michael Mayer（瑞士弗里堡大學(xué)）分享了他的觀點(diǎn)逛揩，即使用聚合物和脂質(zhì)包裹的固態(tài)納米孔對(duì)單個(gè)蛋白質(zhì)淀粉樣蛋白顆粒進(jìn)行高分辨率表征。 SébastienBalme（法國(guó)蒙彼利埃大學(xué)）描述了具有高縱橫比的圓錐形軌跡蝕刻納米孔如何非常適合表征蛋白質(zhì)原纖維的生長(zhǎng)或降解麸俘。

In terms of emerging healthcare applications, it was exciting to see several innovative applications employing solid-state nanopores. Amit Meller (Technion, Israel) shared the latest results from his lab’s work towards using portable nanopore devices for rapid analysis of clinical samples. The technology allows for quantification of RNA expression based on the synthesis and single-molecule counting of gene-specific cDNAs without the need for amplification. Joshua Edel’s and Aleksandar Ivanov’s groups (Imperial College London, UK) demonstrated multiplexed microRNA detection directly in native clinical samples without the need for sample processing using electro-optical nanopore readouts of molecular carriers grafted with molecular beacons. Justin Gooding (University of New South Wales, Australia) adopted a different approach by using magnetic nanoparticle carriers to separate analytes selectively and performed nanopore detection of bioanalytes in whole blood. Vincent Tabard-Cossa (University of Ottawa, Canada) reported on a digital scheme capable of quantifying the concentration of a protein biomarker from serum using DNA nanostructures as proxies for the presence (“1”) or absence (“0”) of the target captured via a magnetic bead-based sandwich immunoassay.

在新興的醫(yī)療保健應(yīng)用方面辩稽，令人興奮的是，看到了一些采用固態(tài)納米孔的創(chuàng)新應(yīng)用从媚。以色列理工學(xué)院的阿米特·梅勒（Amit Meller）分享了他實(shí)驗(yàn)室在使用便攜式納米孔設(shè)備快速分析臨床樣品方面所做工作的最新結(jié)果逞泄。該技術(shù)可以基于基因特異性cDNA的合成和單分子計(jì)數(shù)對(duì)RNA表達(dá)進(jìn)行定量，而無(wú)需進(jìn)行擴(kuò)增拜效。約書(shū)亞·埃德?tīng)枺↗oshua Edel）和Aleksandar Ivanov（英國(guó)倫敦帝國(guó)學(xué)院）的研究小組直接在天然臨床樣品中演示了多重microRNA檢測(cè)喷众，而無(wú)需使用通過(guò)光電信標(biāo)技術(shù)對(duì)接枝有分子信標(biāo)的分子載體進(jìn)行電光納米孔讀數(shù)處理。 Justin Gooding（澳大利亞新南威爾士大學(xué)）采用了另一種方法紧憾，即使用磁性納米顆粒載體選擇性地分離分析物到千，并對(duì)全血中的生物分析物進(jìn)行納米孔檢測(cè)。 Vincent Tabard-Cossa（加拿大渥太華大學(xué)）報(bào)告了一種數(shù)字方案赴穗，該方案能夠使用DNA納米結(jié)構(gòu)作為目標(biāo)存在（“ 1”）或不存在（“ 0”）的代表來(lái)量化血清中蛋白質(zhì)生物標(biāo)志物的濃度憔四。通過(guò)基于磁珠的夾心免疫分析法捕獲。

The majority of these new solid-state nanopore technologies largely rely on molecular probes or carriers based on DNA nanostructures functionalized with recognition sites that enable selective detection of specific analytes. The carrier usually enables more-efficient transport and detection of analytes with a heterogeneous charge that may otherwise be challenging to sense. There are, however, other applications. Ulrich Keyser’s group (University of Cambridge, UK) demonstrated how DNA nanotechnology could be used for information storage and data processing using nanopores. They showed how the density of information could be expanded by increasing the number and the structure of barcodes grafted to a long linear DNA carrier, including 4-way, 6-way and 12-way DNA junctions, that would correspond to logical bits with depths beyond 0 and 1.

這些新的固態(tài)納米孔技術(shù)大部分都依賴于基于具有識(shí)別位點(diǎn)功能的DNA納米結(jié)構(gòu)的分子探針或載體般眉，從而能夠選擇性檢測(cè)特定的分析物了赵。載體通常可以更有效地運(yùn)輸和檢測(cè)帶有異質(zhì)電荷的分析物甸赃，否則這些異質(zhì)電荷很難檢測(cè)柿汛。但是，還有其他應(yīng)用程序辑奈。 Ulrich Keyser的小組（英國(guó)劍橋大學(xué)）展示了如何利用納米孔將DNA納米技術(shù)用于信息存儲(chǔ)和數(shù)據(jù)處理苛茂。他們展示了如何通過(guò)增加嫁接到長(zhǎng)線性DNA載體（包括4向，6向和12向DNA連接點(diǎn)）上的條形碼的數(shù)量和結(jié)構(gòu)來(lái)擴(kuò)展信息密度鸠窗，這些條形碼對(duì)應(yīng)于具有深度的邏輯位超過(guò)0和1妓羊。

While the COVID-19 pandemic forced many of us to distance physically, this virtual meeting’s format can serve as a model for bringing the scientific community closer. A forum for early-career scientists and a training workshop for newcomers took place within the meeting to encourage the next generation of researchers in the nanopore field. The 2020 Nanopore Electrochemistry Meeting was just the start of many more international interactions within this diverse community: following the conference’s success, the Nanopore Weekly Meetings were launched in late October and take place online every Monday. These meetings provide an open platform for researchers worldwide to share new results and exchange ideas on a variety of nanopore-related topics.

盡管COVID-19大流行迫使我們中的許多人物理距離，但這種虛擬會(huì)議的形式可以作為使科學(xué)界更緊密聯(lián)系的典范稍计。會(huì)議期間舉行了一個(gè)針對(duì)早期職業(yè)科學(xué)家的論壇和一個(gè)針對(duì)新移民的培訓(xùn)講習(xí)班躁绸，以鼓勵(lì)納米孔領(lǐng)域的下一代研究人員。 2020年納米孔電化學(xué)會(huì)議只是這個(gè)多元化社區(qū)中更多國(guó)際交流的開(kāi)始：會(huì)議成功舉辦后，10月下旬啟動(dòng)了納米孔每周會(huì)議净刮，并在每個(gè)星期一在線舉行剥哑。這些會(huì)議為世界各地的研究人員提供了一個(gè)開(kāi)放的平臺(tái)，以分享新的結(jié)果并就各種與納米孔有關(guān)的話題交換意見(jiàn)淹父。