Recent Advancements in the SERS-Based Detection of E. coli

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript is very well structured and provides a remarkably comprehensive and systematic explanation that progresses coherently from the fundamental principles of Raman and SERS to their most recent and disruptive applications in E. coli detection. The writing is clear, logically sequenced, and demonstrates a deep understanding of both the physicochemical basis of the techniques and their practical implementation.

One of the greatest strengths is its pedagogical clarity, covering the historical development, enhancement mechanisms (electromagnetic and chemical), and the influence of nanoparticle morphology, laser parameters, and substrate design, not forgetting an up-to-date review of innovative detection methodologies. The section on E. coli quantification is particularly commendable, as it goes far beyond a descriptive account and offers a critical synthesis of diverse approaches.

The tables are also very clear, and summarize particularly valuable and recent information, offering an at-a-glance overview of fabrication methods, enhancement factors, and detection limits that will serve as an excellent reference for different kinds of users.

I have only very minor concerns, as follows:

Although the review convincingly demonstrates the analytical power of SERS for E. coli quantification, it would benefit from a deeper discussion of real-world applicability (e.g., how these laboratory-optimized systems could be adapted for routine industrial monitoring of food and water safety; how they might be standardized and validated for clinical diagnostics)

-Please discuss on cost, robustness, and regulatory acceptance for the real implementation of SERS-based biosensing.

Author Response

Reviewer 1

The manuscript is well structured and provides a remarkably comprehensive and systematic explanation that progresses coherently from the fundamental principles of Raman spectroscopy and SERS to their most recent and impactful applications in E. coli detection. The writing is clear, logically organized, and demonstrates a strong understanding of both the physicochemical foundations of the techniques and their practical implementation.

One of the manuscript’s greatest strengths is its pedagogical clarity. It thoroughly covers the historical development of the field, the enhancement mechanisms involved (both electromagnetic and chemical), and the influence of nanoparticle morphology, laser parameters, and substrate design. It also includes an up‑to‑date review of innovative detection methodologies. The section on E. coli quantification is particularly noteworthy, as it moves beyond descriptive reporting and provides a critical synthesis of diverse analytical approaches.

The tables are clearly presented and summarize valuable and current information. They offer an at‑a‑glance overview of fabrication methods, enhancement factors, and detection limits, making them an excellent reference for a wide range of readers.

I have only a few minor suggestions:

Although the review clearly demonstrates the analytical potential of SERS for E. coli quantification, it would benefit from a deeper discussion of real‑world applicability. For example, how could these laboratory‑optimized systems be adapted for routine industrial monitoring of food and water safety, and how might they be standardized and validated for clinical diagnostics?

Please also include comments on the cost, robustness, and regulatory acceptance relevant to the practical implementation of SERS‑based biosensing.

Response: The authors thank the reviewer for their valuable comments. A dedicated section discussing cost, robustness, and regulatory acceptance has been added to the revised manuscript.

Reviewer 2 Report

Comments and Suggestions for Authors

1. Although abstract is well written, it is suggested to short the abstract.
2. Please check the logic of the third and the forth paragraph in the Introduction section.
3. Introduction: introduction should cover what the authors want to summarize, more information can be added.
4. Section 2.1 can be extended with more information using remarkable achievements.
5. The authors can add a Figure to illustrate the main content of the manuscript.
6. More information can be added in Table 2.
7. A comparison among different solutions is encouraged.
8. A section of challenges and prospects are encouraged to be added in the review. This is critical.
9. The commercial development of these advanced techniques should be discussed
10. A summary of the common patterns of different techniques in Section 5 should be added.

Author Response

Reviewer 2

1. Although the abstract is well written, it is suggested that it be shortened.
   Response: The abstract has been revised and shortened as suggested.
2. Please check the logic of the third and fourth paragraphs in the Introduction section.
   Response: We thank the reviewer for noting this issue. The flow of these paragraphs has been revised for improved clarity and logical progression.
3. The Introduction should clearly convey what the authors aim to summarize; more information can be added.
   Response: We appreciate the reviewer’s input. The Introduction has been revised and condensed to improve clarity and focus, incorporating suggestions from all reviewers.
4. Section 2.1 can be extended with more information, highlighting key achievements.
   Response: Section 2.1 has been expanded to include a concise summary of recent advances in SERS research.
5. The authors can add a figure to illustrate the main content of the manuscript.
   Response: A graphical abstract illustrating the main content has been included.
6. More information can be added to Table 2.
   Response: Table 2 specifically focuses on E. coli Raman spectra, including major peaks, assignments, and interpretations. Additional information on detection methods is already provided in Table 3. Adding more columns to Table 2 would reduce clarity and is not feasible.
7. A comparison among different solutions is encouraged.
   Response: A brief comparison of different solutions has been added to the revised manuscript.
8. A section on challenges and prospects should be included; this is critical.
   Response: A concise section discussing challenges and future prospects has been added.
9. The commercial development of these techniques should be discussed.
   Response: A section on the commercial development of advanced SERS‑based techniques has been added before the summary of Section 5.
10. A summary of the common patterns of the techniques described in Section 5 should be added.
    Response: A summary highlighting common patterns across the techniques in Section 5 has been added.

Reviewer 3 Report

Comments and Suggestions for Authors

This manuscript presents a comprehensive review of surface-enhanced Raman spectroscopy (SERS) for the detection of Escherichia coli (E. coli). The authors first discuss key factors influencing SERS performance, followed by an overview of studies employing various SERS substrates for E. coli detection. Finally, recent advancements integrating SERS with other analytical techniques are introduced. However, several issues outlined below require attention. Therefore, I cannot recommend publication of this manuscript without addressing the following concerns.

 

Line 49: This paragraph can be merged with the preceding one, as they appear to address a similar topic.

Line 89: The term “Moreover” may be inappropriate here and can be replaced with “On the other hand.”

Line 94: The phrase “silver ions” appears unsuitable and should be replaced with “silver.”

Lines 100–103: This section seems redundant and may be combined and reconstructed with the preceding paragraph.

Lines 131–132: Was the silver electrode laser-smoothed? SERS signals typically originate from roughened metal surfaces.

Line 166: This paragraph can be merged with the preceding one, as both discuss related topics.

Line 177: The phrase “In general” seems inappropriate and may be replaced with “On the other hand.”

Line 213: “105-106” -> “10⁵–10⁶”

Line 226: The word “shape” may be omitted, as the paragraph appears to focus solely on the size dependence of SERS.

Lines 255–256: Excitation at shorter wavelengths induces stronger Raman scattering, but not necessarily a higher SERS enhancement factor, which depends on the resonance between the excitation and localized surface plasmon resonance wavelengths.

Line 258: “lower” -> “shorter”

Line 396: The word “Although” may be unnecessary and can be removed.

Lines 414–420: This section appears redundant, as similar content may have been discussed earlier.

Line 430: The abbreviations “NAD” and “FAD” should be written in full upon first mention.

Lines 473–477: As this paragraph serves a conclusive purpose, it may be placed in an inappropriate position and should be relocated.

Table 3: Relevant studies on SERS detection of E. coli via metal ion reduction in or on the cell [Anal. Chem. 2008, 80, 6741; Bull. Chem. Soc. Jpn. 2011, 84, 976] can be cited.

Line 885: The abbreviation “LFA” should be written in full upon first use.

Line 915: The term “Au super particle” should be clearly defined and explained.

Line 935: Relevant references on discrimination using principal component analysis [Anal. Chem. 2004, 76, 40; Analyst 2020, 145, 1236] can be cited.

Reference 39: The bibliographic information appears incomplete. The identity of “N. John” should be verified, as he may not be the editor or author (see https://shop.elsevier.com/books/spectroscopic-methods-for-nanomaterials-characterization/thomas/978-0-323-46140-5).

Reference 103: The page or article number is missing and should be provided.

Author Response

Reviewer 3

This manuscript presents a comprehensive review of surface‑enhanced Raman spectroscopy (SERS) for the detection of Escherichia coli (E. coli). The authors first discuss key factors influencing SERS performance, followed by an overview of various SERS substrates used for E. coli detection. Recent advancements integrating SERS with other analytical techniques are also described. However, several issues must be addressed before the manuscript can be recommended for publication.

Line 49: This paragraph can be merged with the preceding one, as they address similar topics.
Response: The paragraphs have been merged as suggested.

Line 89: The term “Moreover” is inappropriate here; replace it with “On the other hand.”
Response: “Moreover” has been replaced with “On the other hand.”

Line 94: The phrase “silver ions” is unsuitable; replace it with “silver.”
Response: The term has been corrected.

Lines 100–103: This section appears redundant and should be combined with the previous paragraph.
Response: These paragraphs have been combined and restructured.

Lines 131–132: Clarify whether the silver electrode was laser‑smoothed, as SERS signals usually arise from roughened metal surfaces.
Response: We thank the reviewer for noticing this. SERS signals were observed using electrochemically roughened silver electrodes.

Line 166: This paragraph can be merged with the preceding one.
Response: The paragraphs have been merged.

Line 177: Replace “In general” with “On the other hand.”
Response: The change has been made.

Line 213: “105‑106” → “105–106.”
Response: The superscript formatting has been corrected.

Line 226: Remove the word “shape,” as the paragraph focuses solely on size effects.
Response: The word has been removed to avoid confusion.

Lines 255–256: Comment on the relationship between excitation wavelength, Raman scattering, and the SERS enhancement factor.
Response: We have clarified that SERS enhancement scales with the fourth power of the local electromagnetic field and is maximized when the excitation wavelength matches the substrate’s LSPR, especially at shorter wavelengths.

Line 258: Replace “lower” with “shorter.”
Response: Corrected.

Line 396: The word “Although” is unnecessary.
Response: It has been removed.

Lines 414–420: This section is redundant.
Response: The section has been removed.

Line 430: Expand “NAD” and “FAD” upon first mention.
Response: Full forms have been added.

Lines 473–477: This concluding paragraph appears misplaced.
Response: It has been relocated appropriately.

Table 3: Suggested references on E. coli detection via metal ion reduction could be added.
Response: We thank the reviewer; however, Table 3 is focused specifically on E. coli detection studies, and the suggested articles fall outside its defined scope.

Line 885: Expand “LFA” on first mention.
Response: This has been corrected.

Line 915: Define the term “Au super particle.”
Response: A clear definition based on the referenced article has been added.

Line 935: References on discrimination using PCA should be cited.
Response: The suggested reference (Anal. Chem. 2004, 76, 40) has been added.

Reference 39: Bibliographic details appear incomplete; “N. John” is likely incorrect.
Response: Reference 39 has been corrected based on the verified details from the Elsevier book Spectroscopic Methods for Nanomaterials Characterization (Thomas et al., 2017).

Reference 103: Page or article number is missing.
Response: Complete citation details have been added

 

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

1. Response to my comment 2: It seems the authors do not make any improvement. From Line 66, the authors started to introduce spectroscopy, and in Line 74, again. Thus, please refer to my comment 2 again.
2. Response to my comment 3: As the authors have introduced the information in section 2, 3, 4 and 5, some of the information can be introduced in the introduction.
3. Response to my comment 5: which Figure?
4. Response to my comment 6: At least, the instrument information, and the excitation wavelengths should be provided, along with the pretreatment of the samples.
5. Please indicated the line number of the corresponding revisions in the response. It is hard to follow the response. Thus, add the line number of the revision, including the first round of review.

Author Response

Please see attachment

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

The authors may address my comments and revise this manuscript. 

In Line 262, however, it is written as "with substantially greater amplification achievable at shorter wavelengths when..." Excitation at shorter wavelengths induces stronger Raman scattering, but not a higher SERS enhancement factor. Therefore, the phrase "achievable at shorter wavelengths" should be omitted. 

Author Response

Please see attached document

Author Response File: Author Response.pdf