Chemical Images on Fingerprints Revealed with Mass Spectrometry

Round 1

Reviewer 1 Report

The author has shown an innovative method of using TiO2 nanoparticle as a new matrix in MSI for lipids identification in finger marks. TiO2 nanoparticles has an advantage of low background noise that helps in identification of small molecules m/z<500 over conventional matrices.. 

Here, the author mentioned about fatty acid distribution in finger marks for endogenous ones, but fatty acids exist mostly in isomers and also the m/z difference between most of the fatty acids are about 2 Da. How did the author confirm those fatty acids with mass difference of only 2 Da using only MS1 spectrum? It will be challenging due to isotopes overlap. The reviewer is also curious to know the differences between finger marks of two individuals or a pool of individuals. Are those exogenous trace compounds interfere with endogenous ones in negative ion mode MALDI detection?

Author Response

The author has shown an innovative method of using TiO2 nanoparticle as a new matrix in MSI for lipids identification in finger marks. TiO2 nanoparticles has an advantage of low background noise that helps in identification of small molecules m/z<500 over conventional matrices.. 

Here, the author mentioned about fatty acid distribution in finger marks for endogenous ones, but fatty acids exist mostly in isomers and also the m/z difference between most of the fatty acids are about 2 Da. How did the author confirm those fatty acids with mass difference of only 2 Da using only MS1 spectrum? It will be challenging due to isotopes overlap. The reviewer is also curious to know the differences between finger marks of two individuals or a pool of individuals. Are those exogenous trace compounds interfere with endogenous ones in negative ion mode MALDI detection?

 

Thank you for the revision. We reported in Table 2SI a tentative identification with high-resolution mass spectrometry, with ppm accuracy on the identification. We can state only the exact mass of interest, for this reason, we define the identification as “probable”.

Differences between finger marks were identified directly on single finger marks from two different subjects.

Reviewer 2 Report

In general, my opinion is that the article is positive and deserves publication, minor revision is needed before publication.

Please see below some specific questions and comments:

1) When determining the detection limits, nanoparticles were first applied in solution to the target, and then a sample of the standard was applied to them. In real experiments, nanoparticles were applied to the target after contact with the object under study. Is it true that different application methods will not affect the ionization efficiency and, consequently, the sensitivity of the method?

2) Line 121: "Pristine P25 always assists the generation of Na and K molecular adducts " although it is further noted that the processing of nanoparticles makes it possible to obtain other adducts. This is worth reformulating and explaining where cationization comes from. It is also not clear whether this effect is associated with the adsorption of cations on the surface of nanoparticles, or is associated with their hit on the target together with the solution when it is applied? Does the processing time of nanoparticles affect the result? Does the treatment of nanoparticles affect the result of their application for imaging?

3) Fig. 4 - What is the time difference between putting different prints on the target? Could this play a role in connection with the natural degradation (oxidation) of one of the fatty acids?

4) For clarity, please provide the mass spectra for Fig. 4 in the area of each of the fingerprints and in the area of their overlap.

5) There are many works related to the ionization by laser radiation from the surface of titanium oxide (and other metals), which should be mentioned in the introduction and which should be cited (https://pubmed.ncbi.nlm.nih.gov/?term=tio2+nanoparticles+Laser+ionizatoin&sort=date ). It is also necessary to reformulate the conclusions (lines 257-260), paying more attention to the features of the use of nanoparticles, rather than other forms of titanium oxide.

6) The Abstract of the paper should be revised to demonstrate more clearly the results. For example such text as: “Ample evidence has been reported to explain the ionization induced by the TiO2/laser interaction.”. Detection limits and other important specific results should be mentioned in Abstract.

Author Response

In general, my opinion is that the article is positive and deserves publication, minor revision is needed before publication.

Please see below some specific questions and comments:

• When determining the detection limits, nanoparticles were first applied in solution to the target, and then a sample of the standard was applied to them. In real experiments, nanoparticles were applied to the target after contact with the object under study. Is it true that different application methods will not affect the ionization efficiency and, consequently, the sensitivity of the method?

Thank you for the point. Yes, it is correct. We tested sensitivity (and ionization efficiency) and found no differences regarding NP application.

• Line 121: "Pristine P25 always assists the generation of Na and K molecular adducts " although it is further noted that the processing of nanoparticles makes it possible to obtain other adducts. This is worth reformulating and explaining where cationization comes from. It is also not clear whether this effect is associated with the adsorption of cations on the surface of nanoparticles, or is associated with their hit on the target together with the solution when it is applied? Does the processing time of nanoparticles affect the result? Does the treatment of nanoparticles affect the result of their application for imaging?

 

In early experiments, we observed a large cationization of analytes from salts present as residues on NPs and on tissues. For this reason, we decided to prepare NP suspensions in KCl saturated solutions in order to control ionization on a single M+K ion, or at least to favour the potassium cationization. We discussed the process in fig. 4SI, presenting results of doped NP with hard and soft cations, showing an increase of sensitivity with potassium cationization.

A sentence has been modified in Supplementary S1, Lines 64-65in this respect:

“Although the selective cationizations were readily observed, the dispersions were kept under stirring for 72h to facilitate cation exchange and to give more stable results.”

 

• 4 - What is the time difference between putting different prints on the target? Could this play a role in connection with the natural degradation (oxidation) of one of the fatty acids?

Thank you for this point. Unfortunately, we have no information regarding the degradation of fingermarks components, although we obtained similar results even after one day (data not shown). For the image in fig. 4, fingermarks were deposited almost at the same time.

We modified Line 163 in this respect.

• For clarity, please provide the mass spectra for Fig. 4 in the area of each of the fingerprints and in the area of their overlap.

Thank you, we added a new figure 5SI for clarity and added a line in fig. 4 caption.

• There are many works related to the ionization by laser radiation from the surface of titanium oxide (and other metals), which should be mentioned in the introduction and which should be cited (https://pubmed.ncbi.nlm.nih.gov/?term=tio2+nanoparticles+Laser+ionizatoin&sort=date). It is also necessary to reformulate the conclusions (lines 257-260), paying more attention to the features of the use of nanoparticles, rather than other forms of titanium oxide.

Thank you, we added two recent references about TiO2 NP use with a total of 10 references on this topic.

In Conclusions we added a line and references on the generic use of inorganic NPs:

Line 269: “The analysis of small molecules with inorganic NPs, has been described as a powerful alternative to conventional matrices for latent fingerprint [18,41].”

• The Abstract of the paper should be revised to demonstrate more clearly the results. For example, such text as: “Ample evidence has been reported to explain the ionization induced by the TiO2/laser interaction.”. Detection limits and other important specific results should be mentioned in the Abstract.

We agree with the referee. The text “Ample evidence has been reported to explain the ionization induced by the TiO2/laser interaction.” has been removed since it was not directly pertinent with the title.

Sensitivity has been added in line 24 and a sentence in line 28 “… with chemical images that maintain the shape of the objects on the LFM.”

Reviewer 3 Report

I recommend the publication of this manuscript.  Honestly I think that this should have been submitted to a higher profile journal, maybe it was.  The authors described a technique that to my knowledge has not been attempted and that can significantly improve the scope and practical use of MALDI-MS in the field of MS-I with potential applications to areas like forensics, pharmaceutical process monitoring, cosmetics, etc. 

Author Response

I recommend the publication of this manuscript.  Honestly I think that this should have been submitted to a higher profile journal, maybe it was.  The authors described a technique that to my knowledge has not been attempted and that can significantly improve the scope and practical use of MALDI-MS in the field of MS-I with potential applications to areas like forensics, pharmaceutical process monitoring, cosmetics, etc. 

The authors wish to thank you very much for your comments.