Aug 02, 2023
Version 1
sample_prep_serum.nan V.1
Forked from a private protocol
- NAN KB1,
- Mario Uchimiya2,
- John Glushka2,
- Christopher Esselman2,
- Leandro Ponce2,
- Laura Morris2,
- Arthur Edison2
- 1Network for Advanced NMR (NAN);
- 2University of Georgia
- Christopher Esselman: Protocol review;
- Leandro Ponce: Protocol review
Protocol Citation: NAN KB, Mario Uchimiya, John Glushka, Christopher Esselman, Leandro Ponce, Laura Morris, Arthur Edison 2023. sample_prep_serum.nan. protocols.io https://dx.doi.org/10.17504/protocols.io.6qpvr34epvmk/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 02, 2023
Last Modified: August 02, 2023
Protocol Integer ID: 85858
Keywords: serum sample, protein precipitation method, serum, protein, 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and does not constitute legal, medical, clinical, or safety advice, or otherwise; content added to this protocol is not peer reviewed and may not have undergone a formal approval of any kind.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 a protein precipitation method for plasma/serum samples. This protocol was originally proposed by:
Citation
LINK
See also:
Citation
Citation
LINK
Materials
1. Chemicals and reagents
- Phosphate buffer in D2O (100 mM, pH 7.4, 1/3 mM DSS-d6)
- Methanol
2. Equipment
- Calibrated micropipettes (100 μL, 200 μL, and 1000 μL)
- Pippette tips
- 1.5-mL Eppendorf tubes
- 5-mm SampleJet NMR tubes (Bruker)
- Centrifuge
- Vortex mixer
- Speed-vac concentrator
Day-1/2
Thaw samples On ice or at 4 °C
Add 600 µL of 100% cold methanol to 300 µL of samples On ice
- Use 1.5-mL Eppendorf tubes
- Keep methanol cold On ice
Vortex the samples for 00:00:10
10s
Incubate the samples at -20 °C for 00:20:00
20m
Centrifuge the samples at 4 °C at 16000 rcf for 00:30:00
30m
Transfer the supernatants to new 1.5-mL Eppendorf tubes
Dry the samples in a speed-vac concentrator Overnight (time varies depending on the sample condition)
Note
Store dried samples at -80 °C until the following step if needed
Day-2/2
Thaw the samples On ice or in 4 °C
Add 600 µL of the phosphate buffer to each sample
Vortex the samples at 4 °C for 00:10:00
10m
Centrifuge the samples at 4 °C for 00:00:10
10s
Transfer 580 µL to 5-mm NMR tubes
Note
No stickers/labels on the caps and tubes allowed
Citations
Nagana Gowda GA, Raftery D. Quantitating metabolites in protein precipitated serum using NMR spectroscopy.
https://doi.org/10.1021/ac5005103Beckonert O, Keun HC, Ebbels TM, Bundy J, Holmes E, Lindon JC, Nicholson JK. Metabolic profiling, metabolomic and metabonomic procedures for NMR spectroscopy of urine, plasma, serum and tissue extracts.
Nagana Gowda GA, Raftery D. Analysis of Plasma, Serum, and Whole Blood Metabolites Using 1H NMR Spectroscopy.
https://doi.org/10.1007/978-1-4939-9690-2_2