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The Devil is in the Details: Core Adventures in Implementing a New Analytical Method Allis Chien, PhD Director, Stanford University Mass Spectrometry

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Presentation on theme: "The Devil is in the Details: Core Adventures in Implementing a New Analytical Method Allis Chien, PhD Director, Stanford University Mass Spectrometry"— Presentation transcript:

1 The Devil is in the Details: Core Adventures in Implementing a New Analytical Method Allis Chien, PhD Director, Stanford University Mass Spectrometry allis@stanford.edu W ESTERN A SSOCIATION OF C ORE D IRECTORS M EETING S EPTEMBER 12, 2014 …to happy ending? time$$ project goals realitytechnology From challenge…

2 2 SUMS – Stanford University Mass Spectrometry Established 2000 University-wide core resource 8 staff,12 mass specs, 15K samples/year Application areas: Stanford Dean of Research Unit Stanford Cancer Institute Proteomics Shared Resource Vincent Coates Foundation Mass Spectrometry Laboratory Bio-X Mass Spectrometry Core Shared Facility Seely G. Mudd Building Consultation Proteomics Small Molecules Quantitation Drug Discovery Support Open Access Lab Custom Projects

3 Required Administrative Elements GETTING A NEW METHOD OFF THE GROUND new method needtechnologyfunding

4 4 Research project: Acute Respiratory Distress Syndrome More than 190,000 patients develop Acute Respiratory Distress Syndrome (ARDS) every year in the United States, which, even in the modern era, carries an appalling mortality rate of 22-40%.... the etiology of ARDS remains incompletely understood and its clinical course hard to predict. Identifying patients at the highest risk for developing ARDS would enable both early treatment and facilitate enrollment in emerging clinical trials. Project goal: broad, quantitative profiling of common metabolites Challenges: Large number of metabolites Diverse metabolite types Positives: Sharp PI, strong in statistics Access to patients & samples – quality and quantity (200) Understands variability, need for method validation Willing to invest in method development/implementation need

5 “The most extensive metabolite assay in the market. Identification and quantification of more than 180 metabolites from 5 different compound classes” [company brochure] Technology: turnkey method kit for targeted, quantitative analysis of 180 metabolites Tested & published Authentic standards Detailed protocol Pre-developed method Leap of faithProprietaryRegimented tech

6 6 Are these valid solutions?  Core users as a group – build into service rates + Seems logical, for ongoing method development - Usually not equitable  First PI to use – paying for the privilege of being first + No lead time required - Heavy burden for a single PI, often not feasible  First few PIs – share the cost + Spreads out cost - Difficult to predict & coordinate - Slows down process Faculty perspective: “If there is a large upfront cost associated with working with them (i.e. you have to buy some sort of equipment that you don't already own), it would be hard for me to help support it at this point; on the other hand, if it's just a matter of having someone fly over from [Europe], but it's relatively straightforward for your lab to experiment with, I'd be glad to talk about how much that would cost. Certainly doing something with you rather than [Company x] would be great from my perspective for the long-term if the costs can be similar.” Faculty perspective: “If there is a large upfront cost associated with working with them (i.e. you have to buy some sort of equipment that you don't already own), it would be hard for me to help support it at this point; on the other hand, if it's just a matter of having someone fly over from [Europe], but it's relatively straightforward for your lab to experiment with, I'd be glad to talk about how much that would cost. Certainly doing something with you rather than [Company x] would be great from my perspective for the long-term if the costs can be similar.” Funding: Who bears startup costs for new methods? funding

7 7 Seed Grant Program Request verbiage: “A pilot grant program would address several needs simultaneously. Such a program could consist of several small competitive awards per year (e.g. $5-10K each), with the goal of putting together solid mass spec preliminary data for research grant proposals. The funds would certainly support the service center, but unlike a straight subsidy, such a program would raise awareness, increase involvement, and encourage exploration. It would also support staff development by showcasing the collaborative nature of modern mass spectrometry-based research. Similar pilot grant programs have been successful at other universities, and would represent a well-placed investment in future research.” December 2012 Budget request to DoR for Seed Grant Program March 2013 Seed Program approved for FY14 July 2013 Funding Opportunity Announcement issued September 1, 2013 Earliest start date funding Find a link to the FY15 FOA at http://mass-spec.stanford.edu

8 8 ARDS Seed funding proposal Seed grant goals: Supporting the application of existing capabilities to emerging research areas Developing new workflows with existing instrumentation base to advance research Helping PIs generate preliminary data to use in proposals for larger research grants Project aims: 1.Set up metabolite profiling method 2.Verify technical reproducibility via replicates & accuracy via orthogonal measurements 3.Apply method to 200 patient samples need, funding Estimated costs: >>$23K total 1.Method setup – › time: $2,500 › materials: $3,500 2.Verification – › time: $4,000 › materials: $2,000 3.Analysis – › time: TBD › materials: $11,000

9 Administrative stars align – then scientific work begins December 2013 PI project good fit for kit potential funding January 2014 “Background check” discussions with kit provider February 2014 PI applies for seed funding March 2014 $20K seed grant awarded leap of faith preinstall activity April 2014 3-day installation site visit new method needtechnologyfunding

10 Acquired Practical Lessons PUTTING A NEW METHOD INTO PRACTICE new method follow upevaluateprepareinstall Murphy’s Law

11 11 Evaluation: Commercial kit for metabolite quantitation via LC-MS/MS Appealing: Covers nearly 200 metabolites – would be time- and cost-prohibitive to develop from scratch Detailed protocols from sample preparation and MS/MS method setup through data acquisition and analysis Authentic standards & calibration mixes provided Company validated the method, has been used in a number of publications Not so much: Regimented, loss of flexibility batch size – 96 well plate shelf life of months sample type – only for plasma limited dynamic range data analysis workflow Proprietary method, sparse explanations Required >$10K investment before testing Tested & published Authentic standards Detailed protocol Pre-developed method Leap of faithProprietaryRegimented evaluate

12 Don’t assume anything – no detail is too small to ask about and confirm Applications scientist traveling from Europe for the 2-3 day install Kits, software shipped ahead of time Detailed 2-page preinstallation checklist of requirements, including part numbers where relevant: › LC and MS hardware, HPLC column, guard, holder › lab equipment, e.g. evaporator, vortexer, shaker, fume hood, balance › glassware, pipettes › solvents, chemicals › software Discussed item purposes, possible substitutions Didn’t think to ask about software versions… Make a list, check it twice prepare

13 13 More information enables more & better questions Site visit revelations about proprietary methodology Quantitation vs. semi-quantitation Full calibration curve vs. 1 point external calibration LC-MS/MS vs. flow injection analysis (FIA)-MS/MS install installation site visit April 2014 Data collection on replicate set May 2014

14 Corollaries to Murphy’s Law when implementing a new method 1.It will be more complicated than expected 2.It will take more time than expected 3.It will cost more than expected Murphy’s Law

15 Software Requirement: UK English United Kingdom United States Murphy’s Law

16 Follow through with the plan December 2013 Stars align: PI project good fit for kit potential funding January 2014 “Background check” discussions with kit provider February 2014 PI applies for seed funding March 2014 $20K seed grant awarded, leap of faith, preinstall activity April 2014 3-day installation site visit Data collection on replicate set May 2014 Software issues …June 2014… Replicate data released, evaluation in process August 2014 Orthogonal analyses Sept. 2014 200 patient samples October 2014

17 17 Lessons Learned Keep up with new developments Request funding in terms that align with institutional goals Account for all costs, including materials, staff & instrument time Allow for Murphy’s law when generating cost estimates Evaluate feasibility, possible vs. practical Ask questions Pay attention to details Ask more questions Trust and also verify Take the physical location of company personnel into account Build good relationships with support personnel Combine action and patience new method needtechnologyfunding new method follow through evaluateprepareinstall Murphy’s Law

18 18 Acknowledgements Angela Rogers Manuel Kratzke SUMS:  Chris Adams  Ludmila Alexandrova  Karolina Krasinska  Ryan Leib  Theresa McLaughlin  Anna Okumu  Rachel Wu Dean of Research:  Ann Arvin  Sara Bible  John Brauman  Ken Merritt

19 http://mass-spec.stanford.edu Variability Survey & Discussion - A host of factors impact the development of an analytical project. Your survey responses will help stimulate a discussion of variability assessment practices in our community, and how they pertain to the types of experiments that we collectively face every day. Click here to access the survey Click here to access the survey


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