Dear Dr. Misaki,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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We look forward to receiving your revised manuscript.

Kind regards,

Luca Bertolaccini, M.D., Ph.D.

Academic Editor

PLOS ONE

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Additional Editor Comments :

The reviewers have emphasised issues that require a careful and thorough manuscript revision.

No commitment to publication can be made at this point.

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

Reviewer #2: Yes

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

Reviewer #1: Yes

Reviewer #2: No

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3. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.-->

Reviewer #1: No

Reviewer #2: No

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4. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #1: Yes

Reviewer #2: No

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Reviewer #1: I want to start by acknowledging that you have tackled a very practical question in segmentectomy. The overall experimental setup is clinically relevant. Your focus on quantitative ICG behavior is refreshing and useful. I’m sharing the comments below in a constructive spirit:

1. You already present the 5–10 mg/L “sweet spot” and the adequacy of 300 mL/h quite clearly. It may help readers if you end the Abstract and the Discussion with one practical recommendation (something like a clear “this is what we would do clinically based on our findings”).

2. You explain the stepwise design, but it would help to spell out just a bit more transparently how Stage 1 influenced Stage 2. Was patient accrual sequential? And what exactly triggered the choice of conditions in Stage 2?

3. For Stage 1 you say randomization was used, but there’s no detail. Could you clarify how randomization/allocation was actually implemented? Also, for Stage 2/Group F, was every subsequent patient assigned to that group?

4. The in vitro and in vivo parts are both strong, but I think one short sentence connecting concentrations for each group (L/H/F) to the 5–10 mg/L window would make your overall message feel more cohesive.

5. The marking success rate didn’t differ significantly between groups, so the Discussion statement about ICG concentration “significantly affecting” area identification probably needs softening.

6. Several findings hover near significance (e.g., P=0.07) but are described a bit too confidently. I’d suggest framing them more as exploratory/suggestive trends.

7. Since you rely heavily on the DDG 3000 system, consider providing a brief note about its validation/calibration (with a reference if available).

8. Some mechanistic explanations (mixing, bleeding risk, central vs peripheral injection) read a little stronger than what your data directly evaluate. You may just soften the tone or label them as hypotheses.

9. You mention doing a post-hoc power analysis but don’t report the results. Consider providing a quick sentence summarizing it. Also, please make clear where Bonferroni adjustments are applied.

Thank you for sharing your work with me. I genuinely enjoyed reading it.

Reviewer #2: Review of the paper "Evaluation of the effects of indocyanine green administration dose and speed on blood concentration and fluorescence in thoracoscopic segmentectomy". The authors presented a study with two phases: the first in vitro and the second in vivo evaluating the best dose and speed of ICG for identifying the intersegmental plane during segmentectomy. The authors found that the dose of 0.2 mg/kg administered at high speed is associated with a higher concentration of blood ICG and a better visualization of the intersegmental plane. In my opinion, the study is good, but with limited interest because the ICG dose is quite standardized, effective at high dose and without complications or patients risk because the safety profile of the drug is high.

I suggest clarifying some aspects:

1) how was the number of patient selected?

2) I suggest including the type of segmentectomies perfomed

3) I suggest using the interquartile range associated with the median

**********

(what does this mean? ). If published, this will include your full peer review and any attached files.). If published, this will include your full peer review and any attached files.

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Reviewer #1: Yes: Savvas LampridisSavvas Lampridis

Reviewer #2: Yes: Stefano BongiolattiStefano Bongiolatti

**********

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To ensure your figures meet our technical requirements, please review our figure guidelines: https://journals.plos.org/plosone/s/figures

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NAAS will assess whether your figures meet our technical requirements by comparing each figure against our figure specifications.

Dear Dr. Misaki,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please submit your revised manuscript by Apr 01 2026 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org . When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

• A letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols . Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at . Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols ..

We look forward to receiving your revised manuscript.

Kind regards,

Luca Bertolaccini, M.D., Ph.D.

Academic Editor

PLOS One

Journal Requirements:

If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise.

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

Additional Editor Comments:

I would urge the authors to carefully read the remarks of Reviewers and consider these in the revised paper.

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

Reviewer #1: (No Response)

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions??>

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously? -->?>

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.-->

Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #1: Yes

Reviewer #2: Yes

**********

Reviewer #1: Thank you for the revisions and responses. Your manuscript is now much improved, and most of the major issues have been addressed. However, there are a few points that still need your attention:

1. You have correctly clarified that patients were allocated alternately rather than randomly, and I appreciate the correction of the earlier wording. However, I believe this point still needs to be handled more clearly in the manuscript itself. Please describe this as non-random allocation and add a brief acknowledgment of the potential for temporal or secular-trend bias, given the sequential nature of Stage 2 following Stage 1.

2. Although you now state that Bonferroni correction was applied for pairwise comparisons, it still seems unclear to me whether the reported “significant” p-values are adjusted or unadjusted. For example, p-values around 0.03 would not remain significant after Bonferroni correction for three pairwise tests. Please clarify this. Either report Bonferroni-adjusted p-values for pairwise comparisons or distinguish between overall test significance and exploratory/unadjusted pairwise findings.

3. I think there is an inconsistency between the rebuttal and the manuscript regarding how data are summarised. In some places you state that values are presented as median (IQR), while elsewhere the Methods describe medians with min–max. Please standardise this.

4. Your response explaining the distribution of upper versus lower segmentectomies is reasonable, but I think this information should be summarised in the manuscript itself rather than in the response letter or external dataset only (even one sentence in the baseline characteristics or limitations).

Many thanks.

Reviewer #2: This version of the paper has been effectively improved and I do not have any further comment. However, a list of performed segments could be informative for the reader, as I previously suggested.

**********

(what does this mean? ). If published, this will include your full peer review and any attached files.). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our For information about this choice, including consent withdrawal, please see our Privacy Policy .-->

Reviewer #1: Yes: Savvas LampridisSavvas Lampridis

Reviewer #2: Yes: Stefano BongiolattiStefano Bongiolatti

**********

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

To ensure your figures meet our technical requirements, please review our figure guidelines: https://journals.plos.org/plosone/s/figures

You may also use PLOS’s free figure tool, NAAS, to help you prepare publication quality figures: https://journals.plos.org/plosone/s/figures#loc-tools-for-figure-preparation.

NAAS will assess whether your figures meet our technical requirements by comparing each figure against our figure specifications.

Evaluation of the effects of indocyanine green administration dose and speed on blood concentration and fluorescence in thoracoscopic segmentectomy

PONE-D-25-54320R2

Dear Dr. Misaki,

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,

Luca Bertolaccini, M.D., Ph.D.

Academic Editor

PLOS One

Additional Editor Comments (optional):

Reviewers' comments: