PONE-D-21-15429

Salivary proteomics in monitoring the therapeutic response of canine oral melanoma

PLOS ONE

Dear Dr. Suriyaphol,

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Kind regards,

Sompop Bencharit, DDS, MS, PhD, FACP

Academic Editor

PLOS ONE

Additional Editor Comments:

While the editor and reviewer were pleased with the overall work and found the work to be interesting and have potential for publication. We found that there is lack of details in the methods and analysis as well as in the discussion. Please see the detailed comments from the reviewer.

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3.  We noticed you have some minor occurrence of overlapping text with the following previous publication(s), which needs to be addressed:

- https://bmcvetres.biomedcentral.com/articles/10.1186/s12917-020-02550-w

The text that needs to be addressed involves the Materials and Methods sections titled: LC-MS/MS data analysis, Western blot analysis, Verification of expressed protein sequences, and Statistical analyses.

In your revision ensure you cite all your sources (including your own works), and quote or rephrase any duplicated text outside the methods section. Further consideration is dependent on these concerns being addressed.

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Reviewers' comments:

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Comments to the Author

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Reviewer #1: Yes

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: No

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Reviewer #1: Yes

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Reviewer #1: Yes

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5. Review Comments to the Author

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Reviewer #1: This manuscript described the experimental study to utilize the ration of free ubiquitin D (fUBD)and conjugated ubiquitin D (cUBD) in saliva as biomarkers to relate the prognosis oral melanoma in dogs. The authors utilized saliva samples from 9 dogs at different stages of oral melanoma and subjected the samples to tyrpsing digestion and liquid chromatography-tandem mass spectrometry (LC-MS/MS) to verify the presence of the candidate protein including ubiquitin D. Western blot analyses were also used to determine the ration of fUGBD and cUBD.

Overall this review is a very interesting area of research to evaluate the potential features of ratio of fUBD and cUBD in saliva using Western blot analysis and identify the proteins using LC-MS/MS at the different stages of oral melanoma. The authors concluded that There are some questions that remain to be clarified as follows.

1. The figure 4 should be placed before figure 2 and 3 since the identification of UBD was performed before Western blot.

2. Could the author calculate the power analysis? What is the rationale of the sample size?

3. Is there any other way to quantify cUBD and fUBD protein levels besides Western blot analysis?

4. Could the author demonstrate the negative control and positive control in the Western blot analysis?

5. What is the expression profile of normal salivary UBD expression in a normal dog?

6. Would the level of UBD in saliva be affected by the saliva collection time?

7. Please discuss more details on the rationale to focus on the UBD expression in this study. Is there any literature on salivary UBD in other species related to cancer or melanoma? Please discuss.

Thank you very much.

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Reviewer #1: Yes: Phimon Atsawasuwan

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Salivary proteomics in monitoring the therapeutic response of canine oral melanoma

PONE-D-21-15429R1

Dear Dr. Suriyaphol,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

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Kind regards,

Sompop Bencharit, DDS, MS, PhD, FACP

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Thank you for addressing all comments.

Reviewers' comments: