Application of Graphene Oxide as a Biomaterial for the Development of Large-Area Cell Culture Vessels

Round 1

Reviewer 1 Report

Subheading 2.1.1 is duplicated 

2.1.1. Spray coating

GO was purchased...

 2.1.1. Spray coating

For the cell culture tests, fibroblasts...

and in neither case does the paragraph below have any relevance to spray coating. In the first case it describes the GO solution. Clearly this is about the GO source. In which case, rename the subheading and it is important to specify the details of the liquid. Is this really a solution? Is GO soluble in the liquid? I really don't think so. I imagine it is a sol. In which case what is the suspending medium. Is it aqueous? What trace additives are present? Also what is the form of the GO? I presume it is GO nanoparticles. What is their particle size?

In the second case the paragraph describes the cells used. Rename the subheading. The cell description is adequate.

2.2 The Spraying. More details need to be provided. Was the GO "solution" (probably it was a sol) sprayed unaltered from that supplied by "Graphene Supermarket"? If so, all the more important to have details on this "solution". If it was altered in any way before spraying, then details must be provided. If unaltered, it needs to be explicitly stated there. The description in 2.3.1 is confusing in the context: "The spray-coating concentration was set to 0.5 mg/ml, which provided the highest 98 coating efficiency for the amount used in previous studies". What is mg/ml in this context? Does this refer to the supplied "solution"? mg of what? ml of what? It is not sufficient to refer the reader to the literature when a methodological procedure is inadequately described. The literature is used for the introduction and the discussion. In methodology, the materials and processes need to be clearly and unambiguously described so that another scientist can replicated the work.

The results are adequately presented, with some need for a copy edit by a native English speaker. The depth and quality of the data presented is adequate for the purpose.

I have published extensively in the past on the significance of surface roughness on cell attachment. I am satisfied that the authors have presented their findings in this regard satisfactorily, and the novelty of this work is evident. The authors are to be commended on the scientific value and novelty of their findings. 

Author Response

Response to Reviewer 1 Comments

Thank you for your thorough review and valuable comments. We made point-to-point reply to the reviewer’s comments and revisions are reflected in the revised manuscript marked in red.

Point 1: Subheading 2.1.1 is duplicated. In the second case the paragraph describes the cells used. Rename the subheading. The cell description is adequate.

Response 1: ‘Subheading 2.1.1’ is duplicated due to editorial errors. We rewrote the second ‘2.1.1 spray coating’ as ‘2.1.2 cell culture test’ according to the content and the order of paragraph.

Point 2: In the first case it describes the GO solution. Clearly this is about the GO source. In which case, rename the subheading and it is important to specify the details of the liquid. Is this really a solution? Is GO soluble in the liquid? I really don't think so. I imagine it is a sol. In which case what is the suspending medium. Is it aqueous? What trace additives are present? Also what is the form of the GO? I presume it is GO nanoparticles. What is their particle size? 2.2 The Spraying. More details need to be provided. Was the GO "solution" (probably it was a sol) sprayed unaltered from that supplied by "Graphene Supermarket"? If so, all the more important to have details on this "solution". If it was altered in any way before spraying, then details must be provided. If unaltered, it needs to be explicitly stated there. The description in

Response 2: Thank you for your comment. In the case of the GO solution you mentioned, it was purchased at the graphene supermarket and its full name is ultra-highly concentrated single-layer graphene oxide aqueous solution. Flake size is 0.5~5㎛ and thickness is 1 atomic layer(at least 60%).

In general, graphene and graphene oxide are not soluble in water, ethanol, etc. However it is known that GO can be homogeneously dispersed and maintained at least 3 days to 2 weeks. And people use graphene oxide solution as common expression in many papers and products.

Point 3: 2.3.1 is confusing in the context: "The spray-coating concentration was set to 0.5 mg/ml, which provided the highest 98 coating efficiency for the amount used in previous studies". What is mg/ml in this context? Does this refer to the supplied "solution"? mg of what? ml of what? It is not sufficient to refer the reader to the literature when a methodological procedure is inadequately described. The literature is used for the introduction and the discussion. In methodology, the materials and processes need to be clearly and unambiguously described so that another scientist can replicated the work.

Response 3: According to your comment, we added a description for GO solution 0.5 mg/ml in paragraph 2.3.1.

Point 4: The results are adequately presented, with some need for a copy edit by a native English speaker.

Response 4: We received an English language review by a native English speaker through Enago editing institute and referred to this information in the ‘Acknowledgements’.

Reviewer 2 Report

The authors studied the application of graphene oxide as a biomaterial for cell culture vessels. It can be valuable in its own field. However, I have some suggestions:

Major comment:

Some researchers found that graphene oxide was toxic at concentrations of 500 and 1000 μg mL^–1 during a 24 h incubation. Can the author please discuss this point? Why did they choose this material and no other one?

Minor comments

Abstract

please remove the numbers from the abstract.

 introduction: I think the introduction needs to be completely rewritten. It is more about the GO. 

Methods: why does the manuscript have 3 sections with the name 'Spray coating'? What are the differences? the sub-sections of the method section are ambiguous. Please check and re-write.

Please state the details for the  Contact angle and Human Fibroblast cell culture tests. they are so general.

Results

Please add the picture of the angle with the droplet for the Contact angle test.

Conclusion: the conclusion section is so simple! the authors just say 'It was concluded that such a vessel can be developed by optimizing the GO coating method.' please re-write it in a better version.

Author Response

Response to Reviewer 2 Comments

Thank you for your thorough review and valuable comments. We made point-to-point reply to the reviewer’s comments and revisions are reflected in the revised manuscript marked in red.

Point 1: Some researchers found that graphene oxide was toxic at concentrations of 500 and 1000 μg mL–1 during a 24 h incubation. Can the author please discuss this point? Why did they choose this material and no other one?

Response 1: We confirmed through experiments using various cell types such as fibroblast cell, stem cells, and GO concentration of 500 μg mL–1 that the toxicity was not observed when cells were cultured on the spray-coated cell culture plate for more than 24 hours.

Beside GO, many other materials such as molybdenum and titanium can be considered for coating materials. However, those metals are known to be difficult to process and expensive. Graphene oxide is more suitable for cell culture since it is easy to process coating, has biocompatibilty and many functional groups with biological molecules.

Point 2: Please remove the numbers from the abstract.

Response 2: According to your comment, we removed the numbers from the abstract.

Point 3: introduction: I think the introduction needs to be completely rewritten. It is more about the GO.

Response 3: We modified the introduction by adding contents of a GO as a biomaterial.

Point 4: Methods: why does the manuscript have 3 sections with the name 'Spray coating'? What are the differences? the sub-sections of the method section are ambiguous. Please check and re-write.

Response 4: We changed the second ‘2.1.1 spray coating’ to ‘2.1.2 cell culture test’ according to the content and the order of paragraph.

Point 5: Please state the details for the Contact angle and Human Fibroblast cell culture tests. they are so general.

Response 5: According to your comment, we added details on the contact angle in Method. And we changed the title of ‘2.3.7 human fibroblast cells culture test’ to ‘2.3.7 Human bone marrow-derived mesenchymal stem cells (hBM-MSCs) culture test.

Point 6: Please add the picture of the angle with the droplet for the Contact angle test.

Response 6: We added the picture of the angle with the droplet for the Contact angle test in the manuscript.

Point 7: Conclusion: the conclusion section is so simple! the authors just say 'It was concluded that such a vessel can be developed by optimizing the GO coating method.' please re-write it in a better version.

Response 7: Thank you for your comment. We rewrote the conclusion section.

Author Response File: Author Response.docx

Round 2

Reviewer 2 Report

Thanks for the modifications. All modifications are good. However, my two comments about the improving the introduction and the addition of the picture of the angle with the droplet for the contact angle test did not perform proper. the introduction needs improving yet. Please improve it. About the angle with the droplet for the Contact angle test, my mean was your own result's picture. Not schematic picture.

Author Response

Thank you for your thorough review and valuable comments. We made point-to-point reply to the reviewer’s comments and revisions are reflected in the revised manuscript.

Point 1: The improving the introduction

 Response 1: According to your comment, we reorganized the introduction to emphasize the function of GO as a novel biomateial, reinforced the content that is biomaterial application of GO and, added appropriate references.

Point 2: The addition of the picture of the angle with the droplet for the contact angle test.

Response 2: Unfortunately, at the time of the contact angle analysis, the equipment was set to save only the data values ​​calculated by the device. Therefore, we have all the analyzed data values, but no pictures of droplet are saved. So, we added a schematic picture by replacing it.

Author Response File: Author Response.docx