the Creative Commons Attribution 4.0 License.
the Creative Commons Attribution 4.0 License.
3D-printed microcell for protein NMR at high ionic strengths and small sample volumes
Abstract. Standard solution NMR measurements use 5-mm outer diameter (OD) sample tubes that require ca 0.5 mL of solvent to minimize “end effects” on magnetic field homogeneity in the active volume of the sample. Shigemi cells reduce the solvent requirement to ca 0.29 mL. At high ionic strength, or at ultrahigh magnetic fields, smaller OD samples are needed to study samples in conductive, radiofrequency absorbing solvents such as water. We demonstrate an effective and inexpensive alternative for reducing the active sample volume to 0.13 mL by 3D printing of ellipsoidal shaped cells that are inserted into 5-mm OD NMR tubes. Static magnetic susceptibility, χ, of printer resin was measured using a simple slice-selection pulse sequence. We found that the χ of water increases linearly with NaCl concentration, from -9.05 ppm to -8.65 ppm for 0 to 2 M NaCl. The χ of D2O was measured to be -9.01 ppm. The susceptibility difference between the resin (χ= -9.40 ppm) and water can be minimized by paramagnetic doping of the resin. Such doping was found unnecessary for obtaining high quality protein NMR spectra when using ellipsoidal shaped cells that are insensitive to susceptibility mismatching. The microcells offer outstanding RF and good Bo homogeneities. Integrated 600-MHz HSQC signal intensities for the microcell sample in PBS buffer were 6.5±4 % lower than for 0.5 mL of the same protein solution in a regular 5-mm sample tube. The cell is demonstrated for N-acetylated α-synuclein in PBS buffer, and for observing tetramerization of melittin at 2 M NaCl.
Competing interests: AB is a member of the editorial board of Magnetic Resonance.
Publisher's note: Copernicus Publications remains neutral with regard to jurisdictional claims made in the text, published maps, institutional affiliations, or any other geographical representation in this preprint. The responsibility to include appropriate place names lies with the authors.- Preprint
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Status: open (until 20 Apr 2025)
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CEC1: 'Editorial Comment on mr-2025-5', Geoffrey Bodenhausen, 14 Mar 2025
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Our American colleagues may be glad to hear that Copernicus, who produce our journal, will accept payment of the modest Author Page Charges (merely 80 € / page) AFTER their papers are published in their final form, unlike some American publishers who decided to withhold papers that are ready to be published until payment has been received. This may help in these troubled times.
Citation: https://doi.org/10.5194/mr-2025-5-CEC1 -
AC1: 'Reply on CEC1', Ad Bax, 14 Mar 2025
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Thanks Geoffrey. The flexibility of Copernicus is much appreciated and I hope that all our colleagues appreciate that not-for-profit publishing in MR benefits our entire community
Citation: https://doi.org/10.5194/mr-2025-5-AC1
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AC1: 'Reply on CEC1', Ad Bax, 14 Mar 2025
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CC1: 'Comment on mr-2025-5', Tom Barbara, 19 Mar 2025
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This paper is a fine effort in the progression of "small volume NMR", a familiar topic I was introduced to during my days at Varian Associates. In 2009 I wrote up a small review on the history and progress for the eMagRes Journal:
"NMR Probes for Small Sample Volumes" 2009 Thomas M. Barbara https://doi.org/10.1002/9780470034590.emrstm1084
Throughout that history, the issues of dielectric loss, background signals and magnetic susceptibility have always played critical roles. At one time, there was a great effort by a group to etch an approximately elliptical cavity into a glass capillary and this effort reminded me of that work. It would be of some interest to compare this effort with that one (I reference that effort in my article). Background signals in that case are much smaller. This is an aspect of 3D printing materials that can cause headaches for probe builders.Citation: https://doi.org/10.5194/mr-2025-5-CC1 -
CC2: 'Comment on mr-2025-5', Tom Barbara, 19 Mar 2025
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Section 2.1 might perhaps appear rather obscure to a neophyte. For those who are curious, "Cylindrical Demagnetization Fields and Microprobe Design in High Resolution NMR" Journal of Magnetic Resonance A109, 265-269 (1994) will bring satisfaction. It is a real advantage to recognize that magnetization can be viewed as equivalent to a current source whenever there is a gradient or a discontinuity. Back then, it was AOK to use Gaussian units in magnetostatics, but the conversions to SI are actually very simple.
Citation: https://doi.org/10.5194/mr-2025-5-CC2 -
EC1: 'Comment on mr-2025-5', Gottfried Otting, 21 Mar 2025
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Copernicus asks to make data available for the long term. Typically, this is by providing the doi of files stored in a repository (CERN's Zenodo is popular).
Besides the pulse program which is already in the Appendix, it would be nice to make the STL file used for 3D printing available.
Citation: https://doi.org/10.5194/mr-2025-5-EC1 -
AC2: 'Reply on EC1', Ad Bax, 21 Mar 2025
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Our apologies for the omission. The STL file is available at
https://doi.org/10.5281/zenodo.15064675
and this info will be included in the revision
Citation: https://doi.org/10.5194/mr-2025-5-AC2
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AC2: 'Reply on EC1', Ad Bax, 21 Mar 2025
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