Super-Resolution Structured Illumination Microscopy for the Visualization of Interactions between Mitochondria and Lipid Droplets

Round 1

Reviewer 1 Report

Immunostaining and immunoblotting were are cited in the methods, but immunoblot is not shown or cited in the paper. 

In my opinion, the document could be written more clearly. Some sections were hard-to-read. Therefore, I suggest a revision concerning grammar and logic in the “storytelling” of the paper. I read some very clear, ordered, and detailed parts, while others felt like "copied and pasted” with poor attention to the logical flow of the paper. There are also some minor grammatical errors and some wrong tense verbs. These revisions would make the paper easier to read.

The figure legend could be more detailed because sometimes is necessary to go through the paper to find some information about the figure (e.g. Figure 8c colored arrows)

In the discussion part, the authors write “And our results supported the conclusion that mitochondria were responsible for LDs oxidation rather than expansion” but I could not find any link to this topic in the previous pages. I suggest clarifying this better.

Since SIM is wide-spread, I would move figure 1-2-3 in a supplementary section.

I believe that the manuscript by Ting He and collaborators deserves to be published in the MDPI Photonics journal after the above-mentioned revisions.

 

Author Response

Dear Reviewer,

Thank you for your insight and comments concerning our manuscript entitled “Super-Resolution Structured Illumination Microscopy For The Visualization Of Interactions Between Mitochondria And Lipid Droplets”. Those comments are all valuable and very helpful for revising and improving our paper, especially your comments on the details of the article. We have studied comments carefully and have made correction which we hope meet with approval. The article has retained traces of changes for your review. The responds are submitted as an attachment, sincerely thank you and hope to learn more from you.

Author Response File: Author Response.docx

Reviewer 2 Report

Comments for author File: Comments.pdf

Author Response

Dear Reviewer,

Thank you for your insight and comments concerning our manuscript entitled “Super-Resolution Structured Illumination Microscopy For The Visualization Of Interactions Between Mitochondria And Lipid Droplets”. Those comments are all valuable and very helpful for revising and improving our paper, as well as the important guiding significance to our researches, especially for the insertion of references and supplement of experimental details. We have studied comments carefully and have made correction which we hope meet with approval. The article has retained traces of changes for your review. The responds are submitted as an attachment, sincerely thank you and hope to learn more from you.

Author Response File: Author Response.docx

Reviewer 3 Report

He et al. report their development of a 50 fps super-resolution structured illumination microscope (SR-SIM) and their results of imaging interactions between mitochondria and lipid droplets (LDs) in hepatocytes using the microscope. This is an interesting work. The paper is well-written and clear for the readers to understand.

Therefore, I recommend the manuscript be published in Photonics after revisions to address the following points.

Major points:

1. I assume that your conclusion about interactions between mitochondria and (LDs) based on the analysis of the obtained data is correct.

However, I am concerned that the data were 2D. The authors mentioned 3D SR-SIM in the Discussion section. Then, please add descriptions of the following points in association with the thickness of the cell, the spatial resolution of the z-direction, sizes of and shapes of mitochondria and LDs:

Was the xy spatial resolution (132 nm) enough to avoid miscounting the interaction?

Did the overlap between 2D images of mitochondria and LDs that did not make actual physical contact cause misinterpretation?  

2. Figure 6. Please explain why photobleaching for confocal microscopy was more significant than SR-SIM in the main text.

Minor point:

1. Some articles before nouns may be missing. Please check and revise the entire manuscript.

For example,

“stripe pattern” may need an article “a” or may be “stripe patterns” (page 3, line 6).

Each “module” in the sentence may need an article “a” (page 3, line 2 from bottom).

Author Response

Dear Reviewer,

 

Thank you for your insight and comments concerning our manuscript entitled “Super-Resolution Structured Illumination Microscopy For The Visualization Of Interactions Between Mitochondria And Lipid Droplets”. Those comments are all valuable and very helpful for revising and improving our paper, as well as the important guiding significance to our researches, especially for the supplementary discussion of the experimental results. We have studied comments carefully and have made correction which we hope meet with approval. The article has retained traces of changes for your review. The responds are submitted as an attachment, sincerely thank you and hope to learn more from you.

Author Response File: Author Response.docx

Round 2

Reviewer 2 Report

In their revised manuscript, authors He et al. have addressed the recommendations raised in peer review, and in my opinion added many important aspects (robust resolution estimates, implementation choices, etc.) and details (choices for pattern algorithms, polarization maintaining dichroics, etc.) to the manuscript. They also included relevant references to prior works that their research builds on, especially publications that established the 'fastSIM' concept that their microscope is also build on. 

In its current form, I support publication of the manuscript.