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A wearable patch for continuous analysis of thermoregulatory sweat at rest
Low secretion rates and evaporation pose challenges for collecting resting thermoregulatory sweat for non-invasive analysis of body physiology. Here the authors present wearable microfluidics-based patches for continuous sweat monitoring at rest that enable detection of pH, Cl−, and levodopa for dynamic sweat analysis related to routine activities, stress events, hypoglycemia-induced sweating, and Parkinson’s disease.
- Hnin Yin Yin Nyein
- , Mallika Bariya
- & Ali Javey
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A homogeneous split-luciferase assay for rapid and sensitive detection of anti-SARS CoV-2 antibodies
Serological tests are important diagnostic and disease surveillance tools for addressing the COVID-19 pandemic. Here, the authors present a tri-part Nanoluciferase based assay to detect antibodies against SARS-CoV-2.
- Zhong Yao
- , Luka Drecun
- & Igor Stagljar
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2D printed multicellular devices performing digital and analogue computation
Synthetic biology circuits are finding application in a wide range of computational devices, such as contaminant detection. Here, the authors design 2D paper circuits in which the spatial orientation of the cellular components specifies function.
- Sira Mogas-Díez
- , Eva Gonzalez-Flo
- & Javier Macía
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Point-of-care bulk testing for SARS-CoV-2 by combining hybridization capture with improved colorimetric LAMP
Current SARS-CoV-2 diagnostic methods are sensitive yet poorly suited to testing whole communities on a regular basis. Here the authors present Cap-iLAMP that tests gargle lavage samples with an improved colorimetric RT-LAMP.
- Lukas Bokelmann
- , Olaf Nickel
- & Stephan Riesenberg
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Glycan chip based on structure-switchable DNA linker for on-chip biosynthesis of cancer-associated complex glycans
Current methods for on-chip glycan biosynthesis suffer from analysing products, often resulting in poor purity and yield. Here the authors report a glycan chip developed by introducing a pH-responsive i-motif DNA linker to control the immobilization and isolation of glycans on chips.
- Hye Ryoung Heo
- , Kye Il Joo
- & Hyung Joon Cha
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Droplet printing reveals the importance of micron-scale structure for bacterial ecology
The spatial arrangement of bacterial strains and species within microbial communities is considered crucial for their ecology. Here, Krishna Kumar et al. use a droplet-based printing method to arrange different bacterial genotypes across a sub-millimetre array, and show that micron-scale changes in spatial distributions can drive major shifts in ecology.
- Ravinash Krishna Kumar
- , Thomas A. Meiller-Legrand
- & Kevin R. Foster
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An all-to-all approach to the identification of sequence-specific readers for epigenetic DNA modifications on cytosine
Identifying readers of epigenetic marks is a critical step for understanding the role of epigenetic marks in biology. Here, the authors applied DAPPL, an all-to-all approach to profile the interactions between TFs and epigenetic modified DNA libraries.
- Guang Song
- , Guohua Wang
- & Heng Zhu
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A glucose meter interface for point-of-care gene circuit-based diagnostics
Getting synthetic biology circuit-based sensors into field applications is still a challenge. Here the authors combine a circuit sensor with a glucose meter for small analyte and nucleic acid detection.
- Evan Amalfitano
- , Margot Karlikow
- & Keith Pardee
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Analysis of SARS-CoV-2 antibodies in COVID-19 convalescent blood using a coronavirus antigen microarray
COVID-19 diagnosis is commonly performed by PCR testing, however, serologic methods are more accurate and versatile for monitoring disease burden and epidemiology. Here the authors report a protein microarray with antigens from SARS-CoV-2, SARS-CoV, MERS-CoV as well as common human respiratory viruses.
- Rafael R. de Assis
- , Aarti Jain
- & Saahir Khan
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3D projection electrophoresis for single-cell immunoblotting
Single-cell immunoblotting previously separated proteins on a polyacrylamide slab in the xy direction and was limited by throughput and sample consumption. Here the authors adapt the system to separate proteins in the z direction, allowing for closer spacing of sample wells and improved sample consumption.
- Samantha M. Grist
- , Andoni P. Mourdoukoutas
- & Amy E. Herr
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Streamlined inactivation, amplification, and Cas13-based detection of SARS-CoV-2
The COVID-19 pandemic has highlighted the need for user-friendly diagnostic techniques. Here, the authors present SHINE, a streamlined and optimised Cas13-based method with accompanying smartphone app for visual diagnosis.
- Jon Arizti-Sanz
- , Catherine A. Freije
- & Cameron Myhrvold
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An enhanced isothermal amplification assay for viral detection
Current state-of-the-art diagnostics for infectious diseases are sensitive but require extensive equipment. Here the authors develop an enhanced recombinase polymerase amplification reaction for SARS-CoV-2 that allows for inexpensive and rapid testing with minimal equipment.
- Jason Qian
- , Sarah A. Boswell
- & Michael Springer
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Enhancement of trans-cleavage activity of Cas12a with engineered crRNA enables amplified nucleic acid detection
CRISPR-Cas12a based detection systems can be sensitive down to the picomolar range. Here the authors modify the 3′- and 5′-ends of the crRNA and show this enhances trans-cleavage for improved sensitivity.
- Long T. Nguyen
- , Brianna M. Smith
- & Piyush K. Jain
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Ultra-sensitive and rapid detection of nucleic acids and microorganisms in body fluids using single-molecule tethering
Culture-based diagnostic methods for microorganism detection are time-consuming but still the gold standard for conditions such as sepsis. Here the authors present an amplification and purification-free method to detect microorganisms in bodily fluids with high sensitivity: Single MOLecule Tethering (SMOLT).
- Wen-Chih Cheng
- , Troy Horn
- & Alfredo Celedon
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Ultrasensitive and visual detection of SARS-CoV-2 using all-in-one dual CRISPR-Cas12a assay
Rapid and early detection of SARS-CoV-2 will aid intervention to stop disease spread. Here the authors present a one-pot CRISPR-based rapid detection system with visual readout.
- Xiong Ding
- , Kun Yin
- & Changchun Liu
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A role for Biofoundries in rapid development and validation of automated SARS-CoV-2 clinical diagnostics
The SARS-CoV-2 pandemic has created large demand on global testing capability. Here the authors use the London Biofoundry, an automated synthetic biology platform, and develop an open-source virus-like particle to implement high-throughput diagnostics.
- Michael A. Crone
- , Miles Priestman
- & Paul S. Freemont
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Ultrasensitive digital quantification of cytokines and bacteria predicts septic shock outcomes
Ultrasensitive methods for detection of biomarkers for infectious disease are needed for diagnosing, monitoring and targeting treatment. Here the authors develop a digital assay for inflammatory markers, bacterial DNA and antibotic-resistance genes and apply it to characterise asthma patients and predict mortality from septic shock.
- M. Fatih Abasıyanık
- , Krysta Wolfe
- & Savaş Tay
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A CRISPR-Cas9-based reporter system for single-cell detection of extracellular vesicle-mediated functional transfer of RNA
Extracellular vesicles (EV) facilitate intercellular transfer of biological material including RNA, but the regulatory mechanisms for their formation and transfer are incompletely known. Here the authors develop a CRISPR-based reporting system to detect the transfer of guide RNAs and identify genes not previously linked to EV-mediated RNA delivery.
- Olivier G. de Jong
- , Daniel E. Murphy
- & Pieter Vader
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Dissecting splicing decisions and cell-to-cell variability with designed sequence libraries
Alternative splicing is regulated by multiple mechanisms. Here the authors employed designed splice site libraries and massively parallel reporter assays to dissect the regulatory complexity and cell-to-cell variability of splicing decisions and to build accurate predictive models.
- Martin Mikl
- , Amit Hamburg
- & Eran Segal
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The ALFA-tag is a highly versatile tool for nanobody-based bioscience applications
Epitope tags are widely used in various applications, but often lack versatility. Here, the authors introduce a small, alpha helical tag, which is recognized by a high affinity nanobody and can be used in a range of different applications, from protein purification to super-resolution imaging and in vivo detection of proteins.
- Hansjörg Götzke
- , Markus Kilisch
- & Steffen Frey
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A CRISPR-Cas12a-derived biosensing platform for the highly sensitive detection of diverse small molecules
Bacterial allosteric transcription factors can sense and respond to a variety of small molecules. Here the authors present CaT-SMelor which uses Cas12a and allosteric transcription factors to detect small molecules in the nanomolar range.
- Mindong Liang
- , Zilong Li
- & Li-Xin Zhang
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A split fluorescent reporter with rapid and reversible complementation
Monitoring protein-protein interactions via bimolecular fluorescence complementation is often limited by the slow kinetics and irreversibility of the complementation. Here the authors introduce a fluorescent reporter for real-time monitoring of reversible interactions based on complementation and binding of an exogenous chromophore.
- Alison G. Tebo
- & Arnaud Gautier
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Plug-and-play metabolic transducers expand the chemical detection space of cell-free biosensors
The range of chemicals detectable by cell-free systems is still limited. Here the authors combine metabolic cascades with transcription factor networks to detect small molecules in complex environments.
- Peter L. Voyvodic
- , Amir Pandi
- & Jerome Bonnet
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Parallel analysis of tri-molecular biosynthesis with cell identity and function in single cells
Simultaneous quantification of DNA, RNA and protein at the single cell level has not yet been possible. Here the authors introduce a molecular labelling and detection strategy to quantify synthesis of these biomolecules and couple it to transient cell states through parallel quantification of state-dependent biomolecules.
- Samuel C. Kimmey
- , Luciene Borges
- & Sean C. Bendall
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A CRISPR–Cas9-triggered strand displacement amplification method for ultrasensitive DNA detection
Isothermal DNA amplification techniques are useful for diagnostic applications in place of traditional PCR. Here the authors describe CRISDA, which combines CRISPR–Cas9 with strand displacement amplification and exhibits attomolar sensitivity and single-nucleotide specificity in DNA detection.
- Wenhua Zhou
- , Li Hu
- & Xue-Feng Yu
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Microfluidic active loading of single cells enables analysis of complex clinical specimens
Single-cell detection methods are limited by the trade-off between flow rate and measurement precision. Here the authors introduce active loading, an optically triggered microfluidic system to concentrate diluted cell samples, which reduces clogging and decreases processing time in single-cell assays.
- Nicholas L. Calistri
- , Robert J. Kimmerling
- & Scott R. Manalis
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An engineered thermal-shift screen reveals specific lipid preferences of eukaryotic and prokaryotic membrane proteins
Membrane bilayers are made up of a myriad of different lipids that affect membrane proteins, but identifying those specific lipid requirements remains a challenge. Here authors present an engineered thermal-shift screen which reveals specific lipid preferences of eukaryotic and prokaryotic membrane proteins.
- Emmanuel Nji
- , Yurie Chatzikyriakidou
- & David Drew
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Enhanced validation of antibodies for research applications
Five validation pillars have been proposed to verify the specificity of research antibodies. Here the authors screen 6,000 antibodies from the Human Protein Atlas with these methods to provide an antibody validation resource for providers and users.
- Fredrik Edfors
- , Andreas Hober
- & Mathias Uhlen
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rec-YnH enables simultaneous many-by-many detection of direct protein–protein and protein–RNA interactions
Current methods to unveil protein–protein and protein–RNA interactions often involve long, costly procedures. Here the authors present a rapid yeast 2- and 3-hybrid screening pipeline to detect interactions within protein libraries or between protein libraries and RNA fragment pools.
- Jae-Seong Yang
- , Mireia Garriga-Canut
- & Sebastian P. Maurer
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The VAR2CSA malaria protein efficiently retrieves circulating tumor cells in an EpCAM-independent manner
Isolation of circulating tumor cells (CTCs) allows for non-invasive disease monitoring and characterization. Here the authors describe an alternative CTC isolation method based on the ability of the malaria rVAR2 protein to specifically bind oncofetal chondroitin sulfate, which is expressed by all cancer cells
- Mette Ø. Agerbæk
- , Sara R. Bang-Christensen
- & Ali Salanti
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Monitoring the action of redox-directed cancer therapeutics using a human peroxiredoxin-2-based probe
Current sensors for intracellular H2O2 are not sensitive enough to detect all biologically relevant H2O2 fluctuations. Here the authors develop a peroxiredoxin-2-based FRET probe that is sensitive enough to measure changes in H2O2 concentration in response to the cancer therapeutic piperlongumine.
- Troy F. Langford
- , Beijing K. Huang
- & Hadley D. Sikes
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A microfluidics platform for combinatorial drug screening on cancer biopsies
Cancer patients exhibit specific sensitivities toward drug combinations that cannot be easily predicted. Here the authors setup a microfluidic platform that allows testing of multiple drug combinations correctly predicting sensitivity in vivo and they use it on patients biopsies to define effective drugs.
- Federica Eduati
- , Ramesh Utharala
- & Christoph A. Merten
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Performance of the Trioplex real-time RT-PCR assay for detection of Zika, dengue, and chikungunya viruses
The Trioplex real-time RT-PCR assay was developed for detection of Zika virus infections in areas with dengue and chikungunya transmission. Here, Santiago et al. describe the optimization and clinical performance of the assay, showing high sensitivity for detection and differentiation of the three viruses.
- Gilberto A. Santiago
- , Jesús Vázquez
- & Jorge L. Muñoz-Jordan
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Metas-Chip precisely identifies presence of micrometastasis in live biopsy samples by label free approach
Detecting metastatic cells in tumor/lymph node samples of breast cancer patients is extremely important for diagnosis. Here the authors develop a microelectronic biochip that detect the presence of invasive/metastatic cells in unprocessed biopsies and performs better than the current gold standards.
- Mohammad Saeid Nikshoar
- , Mohammad Ali Khayamian
- & Mohammad Abdolahad
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An activity-dependent proximity ligation platform for spatially resolved quantification of active enzymes in single cells
The interrogation of enzyme activity involves the ensemble averaging of many cells, loss of spatial relationships and is often biased to abundant proteins. Here the authors develop activity-dependent proximity ligation to quantify enzyme activity at the cellular and sub-cellular level in relevant biological contexts.
- Gang Li
- , Jeffrey E. Montgomery
- & Raymond E. Moellering
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A signal-amplifiable biochip quantifies extracellular vesicle-associated RNAs for early cancer detection
Extracellular vesicles (EV)-associated RNAs are serum biomarkers potentially exploitable for early cancer diagnosis. Here the authors develop a catalyzed hairpin DNA circuit within a cationic lipid-polymer hybrid nanoparticle that can detect low–level EV-associated RNAs in early stage cancer patients.
- Jiaming Hu
- , Yan Sheng
- & L. James Lee
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Long-term hepatitis B infection in a scalable hepatic co-culture system
The lack of models that mimic hepatitis B virus (HBV) infection in a physiologically relevant context has hampered drug development. Here, Winer et al. establish a self-assembling, primary hepatocyte co-culture system that can be infected with patient-derived HBV without further modifications.
- Benjamin Y. Winer
- , Tiffany S. Huang
- & Alexander Ploss
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Mechanism of SOS PR-domain autoinhibition revealed by single-molecule assays on native protein from lysate
The guanine nucleotide exchange factor SOS is a critical intermediary that transduces receptor tyrosine kinase stimulation into Ras activation. Here the authors develop a single molecule assay in which SOS is captured from raw lysates using Ras-functionalized membrane microarrays and uncover an autoinhibitory mechanism of SOS regulation.
- Young Kwang Lee
- , Shalini T. Low-Nam
- & Jay T. Groves
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Real-time reliable determination of binding kinetics of DNA hybridization using a multi-channel graphene biosensor
Monitoring DNA binding and single-base mismatches accurately in real time is difficult, especially for miniaturized devices. Here the authors report a graphene field-effect transistor array capable of reliably measuring DNA hybridization kinetics and affinity at the picomolar level.
- Shicai Xu
- , Jian Zhan
- & Yaoqi Zhou
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Article
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Real-time quantification of protein expression at the single-cell level via dynamic protein synthesis translocation reporters
Single cells can display large heterogeneity in gene induction. Here, Aymoz et al. present an expression reporter based on protein translocation that can accurately measure both the levels and dynamics of protein synthesis in live single cells with a temporal resolution of less than one minute.
- Delphine Aymoz
- , Victoria Wosika
- & Serge Pelet
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Precision multidimensional assay for high-throughput microRNA drug discovery
Progress in drug discovery can be hampered by a limited exploration of chemical space and the difficulty in assessing the full range of drug candidates’ effects on living cells. Here the authors describe a cell-based assay to distinguish between off-target and specific effects of candidate compounds targeting micro RNAs.
- Benjamin Haefliger
- , Laura Prochazka
- & Yaakov Benenson
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Propagation of oestrogen receptor-positive and oestrogen-responsive normal human breast cells in culture
Culturing normal primary breast cells that express the oestrogen receptor is difficult. Here, the authors isolate oestrogen receptor positive normal breast cells using the cell surface markers CD166 and CD117, and show that the cultures can be repeatedly passaged and retain oestrogen receptor protein expression.
- Agla J. Fridriksdottir
- , Jiyoung Kim
- & Lone Rønnov-Jessen
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Fast and ultrasensitive method for quantitating prion infectivity titre
Bioassays are the standard way to measure prion infectivity titres, but can be time-consuming. In this study, bioassays are compared with a modified version of the protein misfolding cyclic amplification technique with beads (PMCAb), demonstrating that PMCAb can be more precise and faster than bioassays.
- Natallia Makarava
- , Regina Savtchenko
- & Ilia V. Baskakov
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Protein-binding assays in biological liquids using microscale thermophoresis
Protein interactions in biological environments are expected to differ from the situationin vitro. In this study, a thermophoresis-based technique is described that allows the analysis of protein and small-molecule interactions in biological liquids; the work may allow more efficient drug development.
- Christoph J. Wienken
- , Philipp Baaske
- & Stefan Duhr
A rapid, accurate, scalable, and portable testing system for COVID-19 diagnosis