3D printed microchannels for sub-nL NMR spectroscopy

Abstract

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. Here, 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 two-photon polymerization 3D printing. This technique has a resolution better than 1 $μ$m$^3$ and 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 NMR probe consists of an electronic transceiver and a 150 $μ$m diameter excitation/detection microcoil, co-integrated on a single silicon chip of about 1 mm$^2$. 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 2.5x10$^{13}$ spins/Hz$^{1/2}$ on 1H nuclei at 7 T, sufficient to detect highly concentrated endogenous compounds in active volumes down to 100 pL in a measurement time of 3 hours. Spectral resolutions of 0.01 ppm in liquids and of 0.1 ppm in the investigated biological entities are demonstrated. The obtained results indicate a promising route for NMR studies at the single unit level of important sub-nL biological entities, such as living microscopic organisms and eggs of several mammalians, humans included.