Emmanuelle Bardin (Presenter)
Imperial College London
Bio: After graduating in general chemistry from one of the French Grandes Ecoles in 2008, I worked for 4 years as a research engineer at the university and in the industry. My research was mainly applied to the agri-food industry and involved method development using GCMS. In March 2015, I started my PhD in Zoltan Takats’ research group, at Imperial College London. The objective of my project is to develop a quick and non-invasive diagnostic tool which could eventually be adapted to be point-of-care. At the department of Surgery and Cancer, we are working on metabolomic approaches, using ambient MS for the analysis of samples such as skin secretions.
Authorship: Emmanuelle Bardin(1); Frances Bolt(1); Simon Cameron(1); James McKenzie(1); Eric Alton(2); Andrew Bush(2,3); Jane Davies(2,3); Zoltan Takats(1)
(1)Dept of Surgery and Cancer, Imperial College London, London, United Kingdom. (2)National Heart and Lung Institute, Imperial College London, London, United Kingdom. (3)Department of Paediatric Respiratory Medicine, Royal Brompton and Harefield NHS Foundation Trust, London, United Kingdom.
| Short Abstract Sweat contains many chemicals which may provide valuable clinical information. Desorption electrospray ionisation mass spectrometry is a powerful tool for lipid analysis which can detect endogenous lipids in fingermarks. We developed a methodology for the analysis of sweat and examined its utility to assess differences in humans. Principal component analysis demonstrated that lipid related features allowed the differentiation of subjects, gender and diet. The method will be applied to clinical samples in order to examine its potential as a quick and non-invasive diagnostic tool. |
Long Abstract
Here we describe an optimised method for the collection and analysis of secretions from the skin on the bridge of the nose using desorption electrospray ionisation mass spectrometry (DESI-MS). Our overall hypothesis is that individuals secrete a unique lipid profile that may be clinically useful. We demonstrated as a proof of principle that donors, gender and diet could be differentiated. The ultimate aim of the project is to use lipidomics to develop a diagnostic tool for routine clinical use.
Background
Exogenous compounds on the skin surface may originate from the exterior environment or from excretion through sweat of ingested substances, some metabolised, such as drug or food products. Moreover, skin sweat glands secrete endogenous compounds as diversified as inorganic metallic ions, lipids, proteins, vitamins or amino acids [1]. Thus, skin secretions may provide information about a donor’s environment, lifestyle and biology, and represent a huge clinical potential.
The composition of fingermarks in particular has been widely reviewed by forensic scientists where free fatty acids are often identified as the predominant lipids [2]. Mass spectrometry now constitutes the main technique of analysis, especially with the development of mass spectrometry imaging (MSI) [1]. Matrix assisted laser desorption ionization (MALDI) coupled to MSI has been used to investigate the spatial chemical composition of fingermarks [1]. Gender was predicted with 85% accuracy using protein and peptide profiles. Caffeine, fatty acids and amino acid detection has also been reported [3, 4]. This sensitive MALDI technique can analyse a very broad range of molecules but requires sample pre-treatment. The development of desorption electrospray ionisation mass spectrometry (DESI-MS) enabled the ambient analysis of samples in situ. By combining electrospray ionisation with surface desorption, it allows the analysis of solids, adsorbed gases or deposited liquids. Charged solvent droplets generated by a pneumatically assisted electrospray probe are directed towards the sample surface under atmospheric pressure. The solvent dissolves the analytes on the surface and secondary charged microdroplets are desorbed. A subsequent ESI-like process occurs, leading to the formation of gaseous ions of analytes, which are directed towards the MS atmospheric interface [5]. DESI-MS achieved the detection of endogenous lipid compounds, in addition to spiked exogenous drugs or explosives in fingermarks [6].
Preliminary studies showed that fingermarks were subject to environmental contamination including moisturising creams and food, and the results showed poor reproducibility. Therefore, secretions from the bridge of the nose were analysed as these were less prone to contaminating contact. Skincare products were readily eliminated with isopropanol, which did not affect skin secretions. Deposition conditions, substrate and storage were studied and optimised as this can influence sample composition.
Materials and Methods
Secretions were collected on Polysine glass slides (Thermo Fisher scientific) from the skin on the nasal bridges of 14 healthy volunteers of different nationality, gender and diet. Secretions were sampled after cleaning the skin with isopropanol and kept in the freezer at -80C until analysis. The samples were analysed using the Ingeniatrics sprayer as an ion source (Ingeniatrics technologies). The solvent was a mix of methanol/water (95/5 v/v), supplied at a flow rate of 1 uL/min and a nebulising gas pressure of 30psi (N2). The samples were placed on a 3D XYZ moving stage (Prosolia Inc) which also holds the DESI source. The samples are moved by the stage at a speed of 0.1 mm/sec, following a line that crosses the two ink lines and the mark. The analyses are performed with an Exactive Fourier transform MS (Thermo Fisher Scientific), controlled by XCalibur 2.1 software, in positive and negative modes.
Raw files were converted into an mzXML format and processed in a MatLab environment using a customised programme. Pre-processing of the data involved calculating the signal zone median, mass alignment, background subtraction and normalisation. Principal component analysis (PCA) was used for unsupervised analysis of the data set. Analysis of variance (ANOVA) was applied to identify markers with significant contribution. Recursive maximum margin criterion (MMC) was used for supervised classification of the samples. Robustness of the methods was tested through cross validation.
Preliminary data
Fatty acids (200-300 mass over charge (m/z) range) and, at a lower abundance, medium and higher mass lipid species, were detected in negative mode. In positive mode, diglycerides, triglycerides and phospholipids could be detected in the 600-1000 m/z range and dimers of triglycerides were visible in the m/z range 1500 to 1700. The more intense positive ion signal provided better differentiation of the donors. Lipidomic information obtained through DESI-MS allowed the differentiation of each of the subjects and the prediction of their gender and diet was achieved with 90% accuracy.
Conclusions
These first results indicate that nasal skin secretion analysis can be used to investigate lipid-based differences between individuals. Potentially, this is of clinical interest as the human lipidome may yield pertinent information on individual disease states.
Future work will therefore focus on lipid-impacting studies, e.g. diet-based, and identification of differentiating features. We plan to use the method described here for the analysis of nasal skin secretions from patients with a variety of disease in order to examine its utility as a quick and non-invasive diagnosis tool.
References & Acknowledgements:
References
1. Francese, S., et al., Beyond the ridge pattern: multi-informative analysis of latent fingermarks by MALDI mass spectrometry. Analyst, 2013. 138(15): p. 4215-28.
2. Girod, A., R. Ramotowski, and C. Weyermann, Composition of fingermark residue: a qualitative and quantitative review. Forensic Sci Int, 2012. 223(1-3): p. 10-24.
3. Bradshaw, R., et al., Direct detection of blood in fingermarks by MALDI MS profiling and imaging. Sci Justice, 2014. 54(2): p. 110-7.
4. Ferguson, L.S., et al., Direct detection of peptides and small proteins in fingermarks and determination of sex by MALDI mass spectrometry profiling. Analyst, 2012. 137(20): p. 4686-92.
5. Takats, Z., J.M. Wiseman, and R.G. Cooks, Ambient mass spectrometry using desorption electrospray ionization (DESI): instrumentation, mechanisms and applications in forensics, chemistry, and biology. J Mass Spectrom, 2005. 40(10): p. 1261-75.
6. Ifa, D.R., et al., Forensic applications of ambient ionization mass spectrometry. Anal Bioanal Chem, 2009. 394(8): p. 1995-2008.
Supported by the Cystic Fibrosis Trust, Strategic Research Centre for P. aeruginosa
| Description | Y/N | Source |
| Grants | no | |
| Salary | no | |
| Board Member | no | |
| Stock | no | |
| Expenses | no |
IP Royalty: no
| Planning to mention or discuss specific products or technology of the company(ies) listed above: | no |