Dielectric Nanoparticles Coated upon Silver Hollow Nanosphere as an Integrated Design to Reinforce SERS Detection of Trace Ampicillin in Milk Solution

Round 1

Reviewer 1 Report

The manuscript by Han Lee et. al. titled “Dielectric nanoparticles coated upon silver hollow nanosphere as an integrated design to reinforce SERS detection of trace Ampicillin in milk solution” describes a novel approach to creating SERS substrate for detection of ampicillin in milk. The design is based on hollow silver nanospheres with additional coupling to dielectric nanoparticles. Detailed characterization of the manufactured substrates is provided. The authors have attempted to utilize the substrate for the detection of ampicillin in milk using SERS as a readout strategy. The following is the list of major concerns regarding data analysis and representation in this study:

• Page 4, line 135. The prefix is missing. The laser spot could not possibly be 1 m. (there are many instances of missing prefixes in the text)
• SERS EF evaluation is based on a presumption that a uniform distribution is achieved. This is a far-fetched assumption and does not seem to be supported by experimental data. Additionally, it is hard to confirm any calculations in the manuscript without proper prefixes in laser spot diameter, focus length, etc..
• Equation 3 assumes monolayer coverage by R6G with molecules lying flat on the surface, which is clearly not the case considering curved surfaces of the nanoparticles and beads.
• The intensity of Raman increases with increased surface area (section 3.3 and Figure 4): the smaller the beads – the larger the surface area available for R6G to be absorbed. Next, introducing Di-NPs provides even further increase in surface area – thus Raman signal increase. The authors need to clarify this issue.

Minor concerns:

• Page 3, line 110. It is unclear what the following statement means: “It is noted that if there is a film-like Di NPs upon Ag HNS, the incident laser tends to interact with only Di NPs, rather than with the inclusion of Di NPs/Ag interface and Ag HNS, the effect of SERS is presumably reduced. Are there any literature references supporting this statement?
• Page 4, line 124. 10 KV should be 10 kV.
• I would also encourage the authors to consider English editing of the manuscript, especially “Introduction” and “Conclusions”.

Author Response

Dear Editor,

We highly appreciate the reviewers’ comments that help to improve the quality of the revised manuscript. Their addressed questions have been carefully replied.

 Attached please find 4 files: the replies to reviewers, the list of changes, Supporting Data, and the revised manuscript.

Thank you very much for your kind consideration to publish this paper in a regular issue.

Yours sincerely,

Author Response File: Author Response.pdf

Reviewer 2 Report

Lee et al present a substrate for SERS based on hollow silver nanospheres. They use this substrate to detect ampicillin in milk, at low concentrations. The work is interesting and designed sufficiently well. The description is at parts confusing, probably due to limitations in the use of English.

The group has demonstrated in previous manuscripts several SERS substrates, for a variety of applications.

Here, they describe a “silver hollow nano-sphere (Ag HNS) with left inner pore (LIP)” and then coated with dielectrics. The substrate is first characterized with electron microscopy and energy dispersive spectroscopy. The SERS signal is verified first with rhodamine 6g, then with known concentrations of ampicillin in water, and then with a low concentration of ampicillin in milk.

The paper seems to have consistent results, which are for the most part presented in a logical flow. I recommend it for publication after some revisions and language editing.

Itemized comments to authors:

I don’t understand the naming of “LIP”. Left inner pore. Is there a pore in the silver shell? Is it in the shell? What do you mean left? Is there one also on the right? Please reconsider this naming. It is really not clear.

Regarding the experimental procedure: in line 133 you mention that “a fixed amount of rhodamine was dropped onto the test substrate”. What is the volume of sample added to the substrate? Was this left to dry, or scanned in solution? How much later was the spectrum taken? Please explain the whole procedure in more detail, including integration time.

Line 135, please verify the units of the laser diameter.

In line 139 you mention that “Baseline correction and smoothing were performed in conjunction with each other to obtain accurately defined signals.” Please be specific on how these procedures were performed.

In line 145 you mention that “raw milk” was used. Do you mean unpasteurized?
Also, it’s not clear if the sulfuric acid is added to the milk-water solution or to the milk itself.

In line 150 it is stated that “Ampicillin was then added to the as-prepared homogeneous milk solution at a concentration of 0.01 ppm.” Was this the concentration in the milk diluted in water? Or in the initial milk sample? This has implications in the limit of detection reported, and the commercial significance of the report.

Figure 2 where are the red circles?

In line 237, the dielectric constant is stated as 110 k. This cannot be right.

Figure 5 needs units on the y-axis. This will allow the reader to compare intensities in between graphs. It appears that the tick marks do not correspond to the same amount between the different graphs. By adding the units, this graph will be very useful.

 In Figure 6a it would be useful to include the spectrum of the milk solution without addition of ampicillin, to let the reader know if there are any non-specific binding events (as mentioned later in the text).

In figure 6b, the data would be better presented as a bar graph. Connected lines imply time progression and are confusing in this context.

The conclusions are not fully supported by the data, especially the discussion about the mechanism of SERS enhancement. In figure 7 the dielectric islands are proposed to contribute only to the chemical mechanism of enhancement, but in fact can shape the electric field around the dielectric edges creating additional hotspots.

Author Response

Dear Editor,

We highly appreciate the reviewers’ comments that help to improve the quality of the revised manuscript. Their addressed questions have been carefully replied.

Attached please find 4 files: the replies to reviewers, the list of changes, Supporting Data, and the revised manuscript.

Thank you very much for your kind consideration to publish this paper in a regular issue.

Yours sincerely,

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

The authors have addressed my concerns. I would recommend this manuscript for publication.