Aug 17, 2023
sample_prep_urine.nan
Forked from a private protocol
- 1Network for Advanced NMR (NAN);
- 2University of Georgia
- Leandro Ponce: Protocol review;
- Saraa Al Jawad: Protocol review
Protocol Citation: NAN KB, Mario Uchimiya, John Glushka, Leandro Ponce, Saraa Al Jawad, Laura Morris, Arthur Edison 2023. sample_prep_urine.nan. protocols.io https://dx.doi.org/10.17504/protocols.io.j8nlkowkwv5r/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: August 03, 2023
Last Modified: August 17, 2023
Protocol Integer ID: 85925
Keywords: protocol for nmr metabolomic, nmr metabolomic, urine sample, urine, sample
Funders Acknowledgements:
National Science Foundation
Grant ID: 194670
Disclaimer
This protocol is developed and maintained by Network for Advanced NMR (NAN). The protocol content here is for informational purposes only and does not constitute legal, medical, clinical, or safety advice, or otherwise; content added to this protocol is not peer reviewed and may not have undergone a formal approval of any kind. Information presented in this protocol should not substitute for independent professional judgment, advice, diagnosis, or treatment. Any action you take or refrain from taking using or relying upon the information presented here is strictly at your own risk. You agree that neither the Company nor any of the authors, contributors, administrators, or anyone else associated with this protocol, can be held responsible for your use of the information contained in or linked to this protocol or any of our Sites/Apps and Services.
Abstract
This is a modified protocol for NMR metabolomics for urine samples. This method was originally proposed by:
Citation
LINK
See also:
Citation
LINK
Materials
1. Chemicals and reagents
- Phosphate buffer in D2O (1.5 M, pH 7.4, 1.11 mM DSS-d6)
2. Equipment
- Calibrated micropipettes (100 μL, 200 μL, and 1000 μL)
- Pippette tips
- 1.5-mL Eppendorf tubes
- 5-mm SampleJet NMR tubes from Bruker
- Centrifuge
- Vortex mixer
3. Study samples
- Urine samples
Before start
This protocol assumes that the original urine samples have >600 µL.
7m 10s
Day-1/1
Thaw samples On ice or at 4 °C
Centrifuge the samples at 4 °C at 12000 x g for 00:05:00 min
Note
- This step is to remove particles in the samples
- Sample tube and volume specifications vary depending on the study
5m
Add 60 µL of the phosphate buffer to 540 µL of the supernatant for each sample
- Use 1.5-mL Eppendorf tubes
Vortex the samples at 4 °C for 00:02:00
2m
Centrifuge the samples at 4 °C for 00:00:10
Note
This step is to remove samples attached to tube caps, and no centrifugation speed specified
10s
Transfer 590 µL of the supernatant to an NMR tube for each sample
Note
No stickers/labels on the caps and tubes allowed
Citations
Dona AC, Jiménez B, Schäfer H, Humpfer E, Spraul M, Lewis MR, Pearce JT, Holmes E, Lindon JC, Nicholson JK. Precision high-throughput proton NMR spectroscopy of human urine, serum, and plasma for large-scale metabolic phenotyping.
https://doi.org/10.1021/ac5025039Emwas AH, Luchinat C, Turano P, Tenori L, Roy R, Salek RM, Ryan D, Merzaban JS, Kaddurah-Daouk R, Zeri AC, Nagana Gowda GA, Raftery D, Wang Y, Brennan L, Wishart DS. Standardizing the experimental conditions for using urine in NMR-based metabolomic studies with a particular focus on diagnostic studies: a review.
https://link.springer.com/article/10.1007/s11306-014-0746-7