|The manuscript has improved but there are significant shortcomings remaining that have not been addressed or rebutted.|
However, it is encouraging to see that you have engaged with parts of my report despite the fact that you find some comments inappropriate. Unfortunately, this complaint showcases a major issue of the original manuscript and of your rebuttal. You made strong statements and absolute claims and failed to provide the factual basis allowing the interested reader to come to the same conclusions. Numerous statements have been too general and thus factually untrue.
While you clearly state that you consider comment 10 to be inappropriate you fail to state whether you feel that this comment is “hostile or inflammatory” or “libelous or derogatory” in personal comments I make. Nevertheless, I will outline at length why I disagree with you on not showing data and statements you make in response.
According to the Cambridge Dictionary “to refuse” is “to say or show that you are not willing to do, accept, or allow something”. For a fact “data not shown” shows that you are not willing to show this data in this instance which is within the dictionary definition of “refusing”. I hope you will agree that data for central conclusions of a study must be shown otherwise I could publish experimental studies without showing a single piece of data. The common justifications for “data not shown” are confirmatory results, negative results, peripheral results and future results. In the historic context of page limitations there was a point to this. In the age of widespread and unlimited electronic supporting information this really does not apply anymore. The first instance I have criticized mentioned “small” changes with “data not shown”. The problem is that without seeing the data “small” could mean anything from insignificant to substantial and thus, you state a research result that is non-transparent to the reader. In the general context of a science reproducibility crisis in the mainstream media and unlimited supporting material that can be published with most journals more and more publishers are indeed moving away from allowing “data not shown” at all. They do this because there is a growing consensus that full transparency is needed in how results were obtained and conclusions derived and that adhering to this principle will benefit science in the long term.
I am not sure if I understand your point in reply. Are there double measures in that an MR manuscript does not have to be as rigorous as publications in Nature and Science? I do not think this is intended editorial policy. Finally, you might want to reconsider your statement “We are very happy to show all data, although especially nutation experiments are never shown in publications.” This is an absolute (“never”) and insufficiently qualified (“in publications”) statement that is demonstrably wrong. The quantum computing community publish nutation data left, right and center. The editor handling this manuscript has published nutation data, and even in the DEER context these data have been published (e.g., IM-DEER by Clore and coworkers).
1 You evaded my inquiry into the spectroscopic properties of the new model system and the presence of exchange coupling might be relevant even if you are determined to fragmentally publish the characterization of this new compound.
4 There was a clear request for showing Tikhonov regularization results for the mixtures. It is interesting to note that you decided to rather use DeerNet. Nevertheless, this serves the same purpose of a much less heavily parametrized analysis.
6 It is good to see that you can reproduce your data. However, this does not mean that there is no or minimal uncertainty in the extracted parameters. Even if I were to measure several identical and noise-free 2 microsecond DEER I could not reliably extract an 8 nm distance. The request for error or uncertainty estimates remains unanswered. In experimental science a number is only as good as its uncertainty allows. If you do not have a confidence range the number is meaningless.
8 I tend to disagree that giving different names to the same “crosstalk” improves clarity but this will be judged by those who will read and reference this work.
2 and 12 It is unclear how the AWG percentages at 0 dB relate to TWT output. It will be helpful for readers to know if the TWT is operated in saturation at 100% AWG out put or if a lower transmitter level is chosen to linearize the output.
13 You call this an artifact but need a high spin system to observe it. In your own terms this seems more likely to be a signal than an artifact. You also call this spectrometer-specific and even upon request do not provide any evidence that this is indeed a spectrometer specific artifact rather a signal that is not understood.
To repeat myself: It would be very helpful to readers if at least one example per concept (frequency, relaxation and nutation filtering) could be given rather than none at all.
The limitation to 8 nm distances is artificial. There are scenarios where deuteration of the protein is achievable and others where no deuteration is accessible at all (and 8 nm will be beyond reach). If you make absolute and unqualified statements I only need to find a single example to prove you factually incorrect. It is absolutely fine to limit your discussion to a subset of possible experiments but especially the next generation of practitioners will benefit if “too general to be true” statements are avoided.
“We included in the last paragraph of section 1.2 all literature to our knowledge that can be found on NO in conjunction with other spin labels.” Even if this statement was qualified to only consider DEER experiments – which it is not – there are large amounts of literature missed. There is clearly mote that "can be found". https://doi.org/10.1021/ja502615n, https://doi.org/10.1002/chem.201603666, https://doi.org/10.1002/anie.201410396, https://doi.org/10.1007/s00723-010-0181-5, https://doi.org/10.1073/pnas.1200733109, https://doi.org/10.1007/s00723-012-0422-x, https://doi.org/10.1002/anie.201207777, https://doi.org/10.1016/j.jmr.2013.01.002, https://doi.org/10.1039/C8CP06573J to name a few…
It is good to see that hypotheses and outcomes of “comparative analyses” are mostly labelled as such. There is a recent MR discussion paper on Gd-DEER at W-band that may be worth considering in your discussion of the “best” microwave band.
Please check “the Tm values were extracted using MATLAB from the echo decay curves as the time T at which the echo intensity is decayed to 10% of its original value” as this describes something rather different from a (stretched) exponential decay of the form e^(-(2tau/Tm)^x).