7 ^th International Symposium on Enabling Technologies for Life Sciences (ETP)

The seventh in the series of ETP Symposia (see Rapid Communications in Mass Spectrometry 2012, 26 , 1515–1518 for a description and report on the 6 th Symposium) was held at the Metro Toronto Convention Centre, Toronto, Canada, on 30 April to 1 May, 2013, and was chaired by Professor Daniel Figeys, Ottawa Institute for Systems Biology, University of Ottawa. The purpose of these symposia is to convene meetings of scientists and engineers from universities, government laboratories, industry and manufacturers of scientific instrumentation, to discuss novel technologies and methodologies applicable to research in molecular biology. The speakers and the Abstracts of their presentations are listed alphabetically below. The program illustrates some significant changes from earlier versions ( www.etpsymposium.org ) that were devoted almost entirely to proteomics (the 'P' in 'ETP'). In addition to new techniques and applications related to that area, this symposium featured work related to nucleic acid characterization, biomaterials discovery, water disinfection byproducts, and metabolomics, as well as more fundamental developments in mass spectrometry (ionization techniques and quadrupole theory) and sample preparation. The symposium is now more truly international, with speakers from Canada, USA, Europe and China. It is hoped that the wide variety of topics and backgrounds of participants, together with the highly informal setting, will help promote cross‐fertilization of ideas and unforeseen collaborations. Perdita E. Barran , Department of Chemistry, The University of Edinburgh, Edinburgh, UK Resolution of solution structures in the absence of solvent (Invited Talk) Gentle application of nano‐electrospray to proteins buffered in solution to an appropriate pH allows us to use mass spectrometry to interrogate their solution conformations. This can be achieved directly with ion mobility mass spectrometry where we measure the rotationally averaged temperature dependent collision cross section of mass separated ions, or more indirectly with the use of dissociation methods. Data obtained from both methods provides insight to the structure and stability of the protein, and also detail on its interactions, especially when combined with atomistically resolved data from crystallography and/or computational approaches. Data were presented from two systems, the first was a peptide level model of the interaction between the transcription factor c‐MYC and its partner MAX; the second was a detailed structural evaluation of the metamorphic protein Lymphotactin. Ronald Beavis , Beavis Informatics Ltd, Winnipeg, Canada False negatives in proteomics (Invited Talk) Proteomics experiments using tandem mass spectrometry to identify peptides and proteins are prone to various types of false negative identifications that can affect the utility of the information generated from the experiments, as well as their reproducibility. This talk discussed the classification of the most common types of false negatives as well as how they can be detected, remedied or avoided. John D. Brennan, Blake J. Helka and Elna D. Luckham , Department of Chemistry, McMaster University, Hamilton, Canada Imaging MALDI MS/MS of microarrays as a platform for high throughput biomaterials discovery (Invited Talk) This presentation highlighted recent work within the Biointerfaces Institute at McMaster University in the area of high throughput screening of biomaterials, with particular emphasis on the use of imaging MALDI for characterization of biomaterial microarrays. Using robotic handling systems and a combination of contact and noncontact microarray printing, we are able to produce several thousand biomaterials per day with a wide range of chemical compositions. Using sol–gel based materials as an example, the presentation showed the workflow utilized to develop new bioactive sol–gel materials for biosensing and small molecule screening applications. This includes methods to produce several thousand materials very rapidly, tools for rapid screening to identify " hits" that show a desired property (high biological activity, low non‐specific binding, etc.), and further detailed material analysis using a range of imaging methods based on fluorescence, XPS, MALDI‐MS/MS and SPR to fully characterize the properties of biologically active materials. In particular, this presentation emphasized the unique capabilities of MALDI‐MS/MS for evaluating biomaterial properties and the interaction of materials with biological samples. Methods for cataloging, mining and analyzing large datasets within the Database of Biointerface Interactions were also discussed. Rui Chen, H. Zou and D. Figeys , Ontario Institute for Systems Biology, University of Ottawa, Ottawa, Canada Characterization of cell membrane N‐glycosylation with integrated hydrophilic interaction chromatography solid phase extraction and LC/MS/MS Glycosylation of membrane proteins plays important roles in cellular behaviours such as cell‐cell interaction, immunology recognition and cell signalling. Despite their importance the effective extraction of membrane protein, selective isolation of glycopeptides and mass spectrometric characterization of glycosylation are challenging current analytical techniques. In this study, a systematic approach was developed which combined an integrated hydrophilic interaction chromatography solid phase extraction (HILIC SPE) for simultaneous detergent removal and glycopeptides enrichment, and mass spectrometric identification of both protein N‐glycosylation sites and site‐specific glycan structure. The HILIC SPE condition was optimized to enable the use of high concentration of strong detergents, such as SDS and Triton X‐100, to dissolve highly hydrophobic membrane proteins, thus increasing the yield of membrane protein extraction. We illustrated the performance of this approach for the study of membrane protein glycosylation from human embryonic kidney cell lines (HEK 293T). 200 µg total protein digest was processed using this approach, leading to the identification of 813 N‐glycosylation sites from 568 proteins within two experimental replicates. Furthermore, 177 glycopeptides representing 82 N‐glycosides with both glycan composition and peptide sequence were identified by high energy collision dissociation. Andrew Crowell, A. Doucette and S. Rudolph , Department of Chemistry, Dalhousie University, Halifax, NS, Canada. Disposable two‐stage spin cartridges for protein purification in a top‐down proteomics workflow Proper proteome analysis by mass spectrometry depends on upstream sample manipulations, including protein concentration and purification. Cleanup strategies typically rely on chromatography, not only for the relative high recovery and purification efficiency, but also for ease of use through automation. Unfortunately, with chromatographic approaches sample loss is an expected occurrence, especially for high molecular weight and/or hydrophobic (membrane) proteins. By contrast, organic solvent‐assisted protein precipitation is extremely effective at protein purification with minimal sample loss. Quantitative protein recovery is possible, though it is highly dependent on the pipetting skills of the individual. It is desired to create a robust protein precipitation protocol by automating the process, thereby maximizing recovery and purity on a high throughput scale. This presentation introduces a disposable two‐stage centrifugal cartridge, tailored to automate the precipitation process. The ProTrap XG comprises an upper filtration cartridge together with a removable reversed phase cartridge, attaching at its base. Protein samples are mixed with organic solvents in the upper filter chamber, which induces precipitation of proteins. The pellet is retained by the filter, while contaminants in the supernatant are discarded to waste through a brief spin in a desktop centrifuge. Multiple devices can be used simultaneously, improving throughput while maintaining consistency between samples. Following precipitation, protein pellets are re‐solubilized and subject to final purification through the reverse phase cartridge which is re‐attached to the base of the filter. The effectiveness of the

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