- Agilent/Varian cold probe needs 10% of D2O in your protein NMR buffer, 5% of D2O is not working well for the shimming because of its cold probe’s intrinsic design.
- If use a 5mm SHIGEMI tube, at least 320ul of the sample is suggested in order to make the sample length outside the window range of the NMR sample gauge ( window size + 1mm), and further make the Z gradient shimming work well. For the regular 5mm NMR tubes, at least 470~500ul of sample volume is needed.
- When operate “mtune” or “protune”, PATIENTLY wait for “idle” status appearing after every button “clicking”, otherwise it’s easy to block the tuning signal process and freeze the mtune window, and cause to reset the console subsequently!
- During Z gradient shimming process, again, just patiently wait for the macro finishing (about 3minutes), and then “quit” it, otherwise it’s easy to freeze the gradient shimming window too! After you get good Z gradient shimming, you still need to manually shim x, y, and xy… terms.
Use “water” pulse sequence to calibrate proton 90 pulse and then find tof. Use “gNhsqc” pulse sequence to calibrate N15 90 pulse, and use “gChsqc” pulse sequence to calibrate C13 90 pulse for a new protein sample. Re-calibrate the flipback sequence and probefile if needed.
- download the training protocol as your starting point (pdf).
Hongwei updated on Jun. 1, 2016
Edited on Jan. 21, 2014