NMR spectroscopy of single sub-nL ova with inductive ultra-compact single-chip probes

Grisi, Marco; Vincent, Franck; Volpe, Beatrice; Guidetti, Roberto; Harris, Nicola L.; Beck, Armin; Boero, Giovanni

doi:10.1038/srep44670

Cited by 51 publications

(40 citation statements)

References 47 publications

(83 reference statements)

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“…Of particular interest in this picoliter sample-volume regime is the possibility of performing NMR spectroscopy of small molecules and proteins [ 24 ] at the single-cell level. While some work has been done on inductively-detected intracellular NMR with slurries of bacterial cells [ 25 ] and large individual eukaryotic cells such as oocytes [ 26 ], NMR spectroscopy of smaller individual cells has not been achieved. By increasing B0 from 88 millitesla to 1 tesla, both the proton number sensitivity and spectral resolution of an NV ensemble SR sensor can be improved by at least an order of magnitude, which may enable useful single cell NMR.…”

Section: Discussionmentioning

confidence: 99%

High-resolution magnetic resonance spectroscopy using a solid-state spin sensor

Glenn

Bucher

Lee

et al. 2018

Nature

356

454

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Quantum systems that consist of solid-state electronic spins can be sensitive detectors of nuclear magnetic resonance (NMR) signals, particularly from very small samples. For example, nitrogen-vacancy centres in diamond have been used to record NMR signals from nanometre-scale samples, with sensitivity sufficient to detect the magnetic field produced by a single protein. However, the best reported spectral resolution for NMR of molecules using nitrogen-vacancy centres is about 100 hertz. This is insufficient to resolve the key spectral identifiers of molecular structure that are critical to NMR applications in chemistry, structural biology and materials research, such as scalar couplings (which require a resolution of less than ten hertz) and small chemical shifts (which require a resolution of around one part per million of the nuclear Larmor frequency). Conventional, inductively detected NMR can provide the necessary high spectral resolution, but its limited sensitivity typically requires millimetre-scale samples, precluding applications that involve smaller samples, such as picolitre-volume chemical analysis or correlated optical and NMR microscopy. Here we demonstrate a measurement technique that uses a solid-state spin sensor (a magnetometer) consisting of an ensemble of nitrogen-vacancy centres in combination with a narrowband synchronized readout protocol to obtain NMR spectral resolution of about one hertz. We use this technique to observe NMR scalar couplings in a micrometre-scale sample volume of approximately ten picolitres. We also use the ensemble of nitrogen-vacancy centres to apply NMR to thermally polarized nuclear spins and resolve chemical-shift spectra from small molecules. Our technique enables analytical NMR spectroscopy at the scale of single cells.

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Section: Discussionmentioning

confidence: 99%

High-resolution magnetic resonance spectroscopy using a solid-state spin sensor

Glenn

Bucher

Lee

et al. 2018

Nature

356

454

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“…In vivo flow systems were explored in the 1980s but have not become routine due to complications from the intense water peak, broad signals, and low sensitivity . In the last few years, improved water suppression, isotopic enrichment, cryoprobes, microcoils, line narrowing approaches, (removal of susceptibility broadening), and 2D NMR have helped overcome these hurdles, and in vivo NMR is making a resurgence as a powerful technique with unique potential . This tutorial outlines the basics of in vivo NMR for small aquatic organisms and how to build flow systems for researchers interested in exploring the field.…”

Section: Conclusion and Future Prospectsmentioning

confidence: 99%

Flow‐based in vivo NMR spectroscopy of small aquatic organisms

Soong

Mobarhan

Tabatabaei

et al. 2019

Magnetic Reson in Chemistry

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NMR applied to living organisms is arguably the ultimate tool for understanding environmental stress responses and can provide desperately needed information on toxic mechanisms, synergistic effects, sublethal impacts, recovery, and biotransformation of xenobiotics. To perform in vivo NMR spectroscopy, a flow cell system is required to deliver oxygen and food to the organisms while maintaining optimal line shape for NMR spectroscopy. In this tutorial, two such flow cell systems and their constructions are discussed: (a) a single pump high‐volume flow cell design is simple to build and ideal for organisms that do not require feeding (i.e., eggs) and (b) a more advanced low‐volume double pump flow cell design that permits feeding, maintains optimal water height for water suppression, improves locking and shimming, and uses only a small recirculating volume, thus reducing the amount of xenobiotic required for testing. In addition, key experimental aspects including isotopic enrichment, water suppression, and 2D experiments for both 13C enriched and natural abundance organisms are discussed.

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“…[ 55 ] Recently, we reported the use of ultra-compact single-chip NMR probes as convenient tools to deliver state-of-art spin sensitivity for sub-nL volumes. [ 32 , 40 ] Such probes, entirely realized on a single 1 mm 2 complementary-metal-oxide-semiconductor (CMOS) microchip, consist of a multilayer microcoil and a co-integrated low-noise electronic transceiver. Thanks to the achieved sensitivity, we were able to perform the first NMR spectroscopy studies of single untouched sub-nL ova of microorganisms having active volumes down to 0.1 nL.…”

Section: Introductionmentioning

confidence: 99%

3D printed microchannels for sub-nL NMR spectroscopy

et al. 2018

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Nuclear magnetic resonance (NMR) experiments on subnanoliter (sub-nL) volumes are hindered by the limited sensitivity of the detector and the difficulties in positioning and holding such small samples in proximity of the detector. In this work, we report on NMR experiments on liquid and biological entities immersed in liquids having volumes down to 100 pL. These measurements are enabled by the fabrication of high spatial resolution 3D printed microfluidic structures, specifically conceived to guide and confine sub-nL samples in the sub-nL most sensitive volume of a single-chip integrated NMR probe. The microfluidic structures are fabricated using a two-photon polymerization 3D printing technique having a resolution better than 1 μm3. The high spatial resolution 3D printing approach adopted here allows to rapidly fabricate complex microfluidic structures tailored to position, hold, and feed biological samples, with a design that maximizes the NMR signals amplitude and minimizes the static magnetic field inhomogeneities. The layer separating the sample from the microcoil, crucial to exploit the volume of maximum sensitivity of the detector, has a thickness of 10 μm. To demonstrate the potential of this approach, we report NMR experiments on sub-nL intact biological entities in liquid media, specifically ova of the tardigrade Richtersius coronifer and sections of Caenorhabditis elegans nematodes. We show a sensitivity of 2.5x1013 spins/Hz1/2 on 1H nuclei at 7 T, sufficient to detect 6 pmol of 1H nuclei of endogenous compounds in active volumes down to 100 pL and in a measurement time of 3 hours. Spectral resolutions of 0.01 ppm in liquid samples and of 0.1 ppm in the investigated biological entities are also demonstrated. The obtained results may indicate a route for NMR studies at the single unit level of important biological entities having sub-nL volumes, such as living microscopic organisms and eggs of several mammalians, humans included.

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NMR spectroscopy of single sub-nL ova with inductive ultra-compact single-chip probes

Cited by 51 publications

References 47 publications

High-resolution magnetic resonance spectroscopy using a solid-state spin sensor

High-resolution magnetic resonance spectroscopy using a solid-state spin sensor

Flow‐based in vivo NMR spectroscopy of small aquatic organisms

3D printed microchannels for sub-nL NMR spectroscopy

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