Size of Heparin-Imprinted Nanoparticles Reflects the Matched Interactions with the Target Molecule

Round 1

Reviewer 1 Report

Yoshimi et al. studies the size sensitivity of nanoparticles of molecularly imprinted polymers (MIP-NPs) to determine the mechanism of what was previously termed as the “gate effect” by the same author(s). “Gate effect” occurs when nanoparticles immobilized onto a sensing surface (e.g., electrode) swell because of specific interactions with the template, blocking the diffusion pathway of molecular sensing markers to the sensing interface. The authors used nanoparticles of poly(METMAC-co-MBAA-co-AAm) imprinted by heparin as a model and found that the molecularly imprinted polymer nanoparticles swell by matched binding with the template. There are a few pertinent that the authors need to address in this manuscript. While I do not recommend the publication of this manuscript in Sensors in its present state, I believe that it can be reconsidered following the authors’ complete response to the comments below.       

 

Comments

 

The primary objective of the work needs to be clearly presented. While the title gives the impression of emphasis on materials development, the work actually did not focus on developing new materials but rather studying the size sensitivity of MIP-NPs using a procedure modified from previous work by one of the authors.

 

Following the above point, the authors briefly mentioned (e.g., in the abstract) that the work aims understand the mechanism of the “gate effect”. However, it is unclear what new insight is obtained from this work with regard to the mechanism. It is even mentioned in the last paragraph of the Discussion section that “We have yet to obtained definitive information to explain the mechanism of how the matched binding with the template expands the matrix of the MIP…”.The authors need to clarify the most significant, new contribution of this work in terms of mechanistic insight.

 

The Conclusion statement “A nanoparticle of poly(METMAC-co-MBAA-co-AAm) imprinted by heparin swells by matched binding with the template” is rather intuitive and has been described by the author(s) in previous works.  

 

Please include a short discussion on the choice of the “MIP material” (i.e.,      specifically the polymer). How will the use of different MIP material affect the result. Can this be used to reveal greater insght on the size sensitivity of the (MIP-NPs)?

 

Please include a discussion/rationale on the choice of QCM as a characterization method. Can other surface-based characterization tool reveal the same insight? What is the particular benefit of using QCM?  

 

QCM measure the wet mass of adsorbed macromolecules and is therefore prone to hydration effects. How did the authors account for hydration and solvent effects in interpreting the QCM responses?

How many QCM experiments are performed? For Figure 3, the reviewer recommends the authors to accompany the real-time responses of the QCM signal with a plot of the maximum frequency shift, with error bars included. This will provide a better indication of the reliability of the QCM data. 

Author Response

Thank you very much for your comments. Please find the attached file.

Author Response File: Author Response.docx

Reviewer 2 Report

In this article, the authors investigate a potential mechanism that might explain previously obtained results. In this way, the authors demonstrate that the selectivity of their electrochemical sensor does not necessarily mean that the analogue does not bind, as they demonstrate the opposite with QCM. The authors provide a potential explanation for this apparent discrepancy by studying the effect of target and analogue binding on the change in diameter in the nanoparticles. This is coupled to an elegant explanation that does not only provide a possible explanation for the observed selectivity but also the difference in behavior observed on grafted electrodes on the one hand and carbon-paste electrodes on the other hand. As this study is interesting for and relatively novel within the field of MIP-based sensing, I recommend publishing the article upon addressing following issues:

 

1.     I think the abstract is a bit confusing. When I read that the size sensitivity would be investigated, I was under the assumption that the size of the nanoparticles would be varied and its effect on the MIPs performance would be analyzed. Rather the authors investigate how binding of target and analogue changes the diameter.

2.     The introduction is lacking a bit of background in terms of references. The authors only included 19 references of which a substantial amount of self-references. I would add some more general references if you explain the importance of a MIP and some more specific on MIP-based sensors using different readout mechanisms. I would include at least impedance-based sensing as well as thermal and colorimetric detection. 

3.     Is there an explanation for not being able to obtain NIP particles?

4.     Is there any reason for not studying LMWH-coated quartz crystals? An analysis of their response towards UFH and LMWH imprinted MIPs could be very interesting. Although I realize that this experiment is just done to illustrate that not seeing an electrochemical response does not mean the CSC does not bind.

5.     The conclusion is a bit underwhelming. It is a bit too concise.  

 

Minor isssues

1.     The sentence "The high selectivity of the MIP-NPs was consistent..." in the intro is quite long. I suggest that you remove "where the redox current was sensitive to the concentration of heparin, but insensitive to 25 CSC.". You already said that in the previous sentence and then say it is similar now so that should suffice. 

2.     The "from" on line 37 page 1 seems redundant.

3.     It is a bit weird to refer to a paper as “developed by one of us and colleagues”

4.     The font size decreases in the methods section, which is not uncommon but stays small, increases again and then decreases one more time in the results section. The authors should keep this constant.

5.     In the result section, I would use subheadings to make the different parts more clear and I would discuss the results figure per figure. Additionally, I would add legends to the figures instead of putting the explanation in the caption.

Author Response

Thank you very much for your kind advises. Please find the attached file.

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

Overall, the revisions are fine. For Figure 3, the author strongly recommends the accompaniment of the real-time responses of the QCM signal with a plot of the maximum frequency shift, with error bars included. This will make Figure 3 more complete.  

Author Response

Dear Reviewer 1

Thank you very much. According to your advise, We added the Fig. 3D