Methods and microfabrication techniques for subnanoliter magnetic resonance spectroscopy

Nuclear magnetic resonance (NMR), electron paramagnetic resonance (EPR) and ferromagnetic resonance (FMR) spectroscopy can open up the possibility of studying many scientifically and biologically relevant samples at the µm and sub-µm scale. Examples of volume-limited systems include numerous species of microorganisms, mammalian zygotes, the majority of cells, proteins limited in growth, micro and nanostructured devices for the analysis of spin dynamics at the sub-µm scale. These volume-limited samples cannot be addressed by commercially available inductive spectrometers due to their constraint in sensitivity. It was previously proposed in our group that CMOS technology can be used to realize miniaturized high sensitivity inductive detection systems, having spin sensitivities at least two orders of magnitude greater than the commercially available spectrometers. During my PhD work, I developed methods and microfabrication techniques to perform NMR, EPR and FMR spectroscopy at the µm and sub-µm scale by using high sensitivity single chip CMOS detectors, previously proposed in our group. Microfluidic systems for the non-invasive handling of liquid samples and biological entities immersed in liquids are realized and integrated with the CMOS single chip NMR and EPR detectors. Microfluidic channels are fabricated via conventional microfabrication techniques and via two-photon polymerization, a 3D printing technique with a lateral resolution of 300 nm. The 3D printing technique is found to be an exceptional solution for NMR applications. Due to the flexibility in the design of the microfluidic systems, it is possible to reduce the magnetic field non-uniformieties with a consequent improval in spectral resolution. Spectral resolutions down to 0.007 ppm are reported for liquids having sample volumes of 100 pL. For the first time, NMR studies on intact biological entities submerged in liquid media of choice are performed, using 3D printed microfluidic systems. A spin sensitivity of 2.5·1013 spins/¿Hz is shown, sufficient to detect highly concentrated endogenous compounds in active volumes down to 100 pL with measurement times down to 3 h. EPR measurements on subnanoliter liquids and frozen solutions are reported, by the combination of commercially available capillaries and EPR single chip detectors. This is a first but important step towards the study of biological samples, whose paramagnetic ions have relaxation times too short to be measured at room temperature. Moreover, a novel method for the sensing of magnetic microbeads is presented, which is based on the detection of the change of susceptibility in FMR condition by the CMOS integrated detector. Due to the frequency and field dependence of the susceptibility, the detected variation is 20 times greater than the change in magnetization measured in static conditions by other approaches. The proposed detection scheme allows for single bead sensitivity over an active area of about 5·104 µm2. Lastly, sub-µm scale FMR detection capabilities of the single chip CMOS detector are shown by experiments on nanopatterned single permalloy (80% Ni - 20% Fe) and YIG (Yittrium Iron Garnet) dots. The combination of high sensitivity and large active area is seen as a considerable advantage with respect to other FMR detection methods, which suffer either from sensitivity or from reduced active area.

Novel Sample Formulations for Pure and Persistent Hyperpolarized Solutions via Dissolution Dynamic Nuclear Polarization

Basile Vuichoud

Nuclear Magnetic Resonance (NMR) spectroscopy allows one to study and analyze the structure, motions and interactions of a broad variety of molecules. However, this technique has a major inconvenience: its low sensitivity, which therefore often results in the use of highly concentrated samples in order to observe tiny signals. As recently as 2003, a new technique known as dissolution-DNP or d-DNP was invented by Ardenkjaer-Larsen et al. to overcome this drawback and to get more intense NMR signals in solution with enhancements factors larger than 10'000. This method consists essentially in mixing paramagnetic species (radicals) with samples containing the metabolites to be analyzed. These mixtures are then rapidly frozen at very low temperatures (T = 1.2 â 4.2 K) in liquid helium and, by applying a proper microwave irradiation, the polarization of the electrons can be transferred to nuclei such as 1H or 13C. This allows one to build up and store the enhanced magnetization of these nuclei and then to dissolve the samples by injecting a superheated solvent via a dissolution stick. The resulting hyperpolarized solution can be finally transferred to an NMR spectrometer where the signals can be recorded. In the first two chapters of this thesis, the principles of NMR are introduced and the theory of DNP is explained in some detail. The dissolution equipment used in our laboratory and its different parts are shown, in particular the DNP polarizer where the transfer of polarization from electrons to nuclei occurs, the microwave source that is connected to the polarizer, and the dissolution system itself, which comprises the dissolution stick and the dissolution transfer line. Another chapter is dedicated to the optimization of our DNP setup in order to achieve the highest possible polarization before the dissolution process. Several radicals are tested under carefully controlled conditions to identify the best suited for our DNP system. Furthermore, the modulation of the microwave frequency has been optimized in order to enhance the polarization transfer. Finally, a number of dissolution experiments are presented that relate to different projects that have been carried out in the course of this thesis. The extent of the polarization can be determined accurately by looking directly at the hyperpolarized NMR spectrum. Filterable polymers containing suitable radical moieties have been synthesized in order to obtain pure hyperpolarized solutions. A final chapter outlines future applications and projects that can benefit from dissolution Dynamic Nuclear Polarization.

EPFL2017

Chemical Developments in DNP-NMR

Pascal Miéville

The Nuclear Magnetic Resonance (NMR) is an extraordinary tool to understand molecular structures. Its main limitation resides in its relatively low sensitivity when compared with other physico-chemical methods such as Mass Spectrometry (MS) or Optical Spectroscopy. Dynamic Nuclear Polarization (DNP), invented in the 50's but only widely applied in the last ten years, allows, by transferring the high polarization of paramagnetic centers to surrounding nuclei at low temperature, to enhance NMR signals by several orders of magnitude. This technique can be applied to solids at 100 K using Magic Angle Spinning (DNP-MAS) or in liquid phase through Dissolution-DNP as described by Ardenkjaer-Larsen in 2003. The efficiency of the polarization transfer is strongly dependent on the nature of the paramagnetic centers and on the way they are mixed with the target molecules. Part of this work will consist in the study of new ways to integrate paramagnetic centers into the matrix and also to limit their negative effects on NMR signals in liquid phase after dissolution. Because DNP-NMR has reached a stage where it should become a standard chemical method, a second objective of this thesis is to propose new chemical applications such as kinetic measurements that may be of interest in other domain of chemistry.

EPFL2011

A single chip electron spin resonance detector based on a single high electron mobility transistor

Giovanni Boero, Alessandro Valentino Matheoud, Nergiz Sahin Solmaz

Single-chip microwave oscillators are promising devices for inductive electron spin resonance spectroscopy (ESR) experiments on nanoliter and subnanoliter samples. Two major problems of the previously reported designs were the large minimum microwave magnetic field (0.1 mT to 0.7 mT) and large power consumption (0.5 mW to 200 mW), severely limiting their use for the investigation of samples having long relaxation times and for operation at low temperatures. Here we report on the design and characterization of a single-chip ESR detector operating with a microwave magnetic field and a power consumption orders of magnitude lower compared with previous designs. These significant improvements are mainly due to the use of a high electron mobility transistor (HEMT) technology instead of a complementary metal-oxide-semiconductor (CMOS) technology. The realized single-chip ESR detector, which operates at 11.2 GHz, consists of an LC Colpitts oscillator realized with a single high-electron mobility transistor and a co-integrated single turn planar coil having a diameter of 440 μm. The realized detector operates from 300 K down to 1.4 K, at least. Its minimum microwave magnetic field is 0.4 μT at 300 K and 0.06 μT at 1.4 K, whereas its power consumption is 90 μW at 300 K and 4 μW at 1.4 K, respectively. The experimental spin sensitivity on a sensitive volume of about 30 nL, as measured with a single crystal of α,γ-bisdiphenylene-β-phenylallyl (BDPA)/benzene complex, is of 8×10^(10) spins/Hz^(1/2) at 300 K and 2×10^(9) spins/Hz^(1/2) at 10 K, respectively. In a volume of about 100 pL, located in proximity to the coil wire, the spin sensitivity improves by two orders of magnitude.

2018

Novel Sample Formulations for Pure and Persistent Hyperpolarized Solutions via Dissolution Dynamic Nuclear Polarization

Basile Vuichoud

EPFL2017