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CURRICULUM

VITAE

Nathan A. Siegfried

Saladax Biomedical, Inc. Home (preferred)

*** ******** **. **** ********** Rd

Bethlehem, PA 18015 Allentown, PA 18104

Desk: 484-***-**** Tel: 610-***-****

adf6vj@r.postjobfree.com adf6vj@r.postjobfree.com

Experience:

April 2017 – Present Principal Scientist, Saladax Biomedical Inc.

• Project leader in R&D; design, develop, and validate immunoassay medical devices for clinical sample analysis

o Design, troubleshoot assay components, parameters to function with human serum/plasma/blood, urine o Operated in a GMP/GLP environment, created controlled documents, design input, output content o Analyzed product stability, matrix effects, verified quality of components and de-risked processes o Contribute to regulatory (CE mark, FDA) submissions, product development process

Project leader on medical device recently granted by the US FDA o Manage one direct report employee

o Technology transfer to production

o Monitor psychiatric scientific literature and produce marketing literature o Collaborate with psychiatrists, psychologists, and mental health advocacy groups o Represent Saladax at conferences; American Psychiatric Assn, AACC June 2014 – Dec 2016 Senior Scientist, The University of North Carolina at Chapel Hill Professor J. Michael Ramsey, Department of Chemistry, Biomedical Engineering, Lineberger Cancer Center

• Main project: Develop microfluidic molecular diagnostic assay platform for commercialization o Design, troubleshoot qPCR, immunoPCR, ELISA, SiMOA assays in a microfluidic system o Characterize fluorophore influence on PCR, ELISA in microwell arrays via microscopy o Supervise method transfer for downstream application on custom instrument o Develop custom formulations for in-chip storage of reagents o Lead design of sample processing strategies to isolate targets in blood, sputum, bodily fluids o Supervise five postdoctoral fellows, two graduate students, and one technician

• Secondary project: Develop automated process for stem cell selections in microwells o Advise graduate student in assay development performed on cells in microwell devices Feb-June 2014 Consultant, The University of North Carolina at Chapel Hill Professor Alain Laederach, Department of Biology

• Supervised transfer of RNA analytical methods and protocols I developed as a postdoctoral fellow

• Facilitated and supervised the work of three postdoctoral associates and one graduate student

• Redesigned laboratory space and workflow to support bioanalytical experiments (RNA structure probing) 2009-2014 Postdoctoral Fellow, The University of North Carolina at Chapel Hill Advisor: Professor Kevin M. Weeks

• Pioneered novel, enzyme-mediated analytical method for RNA structure analysis using NGS o Developed and optimized unique reverse transcriptase enzymatic activity o Obtained intellectual property for this technology, currently being commercialized (Ribometrix)

• Structurally characterized entire HIV RNA genome, designed mutations to disrupt viral function

• Cloned, expressed, and purified recombinant proteins used in analytical assays

• Human tissue culture and virology; assessed HIV infectivity with various virus strains o Trained in Biosafety level 2 and 2 Plus protocols

• Awarded two competitive research fellowships

2003 – 2009 Graduate Student, The Pennsylvania State University Advisor: Professor Philip C. Bevilacqua

Focus: RNA catalysis via proton exchange: RNA folding cooperativity and protonation states

• Analyzed biophysics of RNA using many traditional biophysical analytical methods

• Characterized folding cooperativity and effects on nucleobase pKa values in DNA and RNA

• Used modified nucleobases to dissect folding thermodynamics

• Taught senior-level analytical lab (HPLC, GS-MS, FT-IR, CE and electrochemistry) Page 2 of 3

Education

Ph.D., Chemistry

The Pennsylvania State University, University Park, Pennsylvania, Focus on RNA and DNA biophysics B.S., Chemistry (Minor: Biology)

Moravian College, Bethlehem, Pennsylvania

Magna Cum Laude, G.P.A.: 3.77, Honors in chemistry, Research in organic mechanisms Selected Publications:

Siegfried N.A.; Busan S; Rice G.M.; Nelson J.A.E; Weeks, KM. “RNA motif discovery by SHAPE and mutational profiling (SHAPE-MaP).” Nat Methods, Sep; 11(9), 959-969 (2014). Siegfried N.A.*; Kierzek R.; Bevilacqua P.C. “Role of Unsatisfied Hydrogen Bond Acceptors in RNA Energetics and Specificity.” Journal of the American Chemical Society, 132(15), 5342-5344 (2010). *Corresponding author Patent: Weeks, K.; Siegfried, N.A.; et. al. 2014, Detection of Chemical Modifications in Nucleic Acids. US Patent Application/ PCT 15/027,813

Publications:

Buckley, T., Kitchen, C., Vyas, G., Siegfried, N., Tefera, E., Chen, S., DiPaula, B., Kelly, D. “Comparison of novel immunoassay with liquid chromatography/tandem mass spectrometry (LC-MS/MS) for therapeutic drug monitoring of clozapine.” In press, Therapeutic Drug Monitoring, March 2020. Henley, W., Siegfried N.A., Ramsey, J.M. “Spatially Isolated Reactions in a Complex Array: Using Magnetic Beads to Purify and Quantify Nucleic Acids with Digital and Quantitative Real-time PCR in Thousands of Parallel Microwells.” In press, Lab On A Chip, March 2020.

Gaiteri, J.C.; Henley, W.H.; Siegfried, N.A.; Linz, T.H.; Ramsey, J.M. “Use of ice-nucleating proteins for enhanced performance of freeze-thaw valves in microfluidic devices.” Analytical Chemistry, 89, 11 (2017). Smola, M.J.; Rice, G.M.; Siegfried, N.A.; Weeks, K.M. “Selective 2ˈ-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP): direct, versatile, and accurate RNA structure analysis.” Nat Protocols, 10, 1634-1669 (2015).

Mauger D.M.; Siegfried N.A.*; Weeks K.M. “The Genetic Code as Expressed Through Relationships Between mRNA Structure and Protein Function.” FEBS Letters, 587, 1683-1694 (2013). *Co-first author Chadalavada, D.M.; Cerrone-Szakal, A.L.; Wilcox, J.L.; Siegfried, N.A.; Bevilacqua, P.C. “Mechanistic Analysis of the Hepatitis Delta Virus (HDV) Ribozyme: Methods for RNA Preparation, Structure Mapping, Solvent Isotope Effects, and Co-transcriptional Cleavage.” Methods in Molecular Biology, 848, 21-40, (2012). Steen, K.A.; Siegfried, N.A.; Weeks, K.M. “Selective 2ʹ-Hydroxyl Acylation analyzed by Protection from Exoribonuclease (RNase-detected SHAPE): direct analysis of covalent adducts and of nucleotide flexibility in RNA." Nature Protocols, 6, 1683-1694 (2011).

Siegfried N. A.; O’Hare B.; Bevilacqua P.C. “Driving Forces for Nucleic Acid pKa Shifting in an A C Wobble: Effects of Helix Position, Temperature, and Ionic Strength.” Biochemistry, 49(15), 3225-32236 (2010). Siegfried, N. A.; Bevilacqua, P. C. “Thinking Inside the Box: Designing, Implementing, and Interpreting Thermodynamic Cycles to Dissect Cooperativity in RNA and DNA Folding.” Methods in Enzymology, 455, 365-393

(2009).

Cerrone-Szakal A. L.; Siegfried N. A.; Bevilacqua P. C. “Mechanistic Characterization of the HDV Genomic Ribozyme: Solvent Isotope Effects and Proton Inventories in the Absence of Divalent Metal Ions Support C75 as the General Acid.” Journal of the American Chemical Society, 130(44), 145**-***** (2008). Siegfried, N. A.; Metzger, S. L.; Bevilacqua, P.C. “Folding Cooperativity in RNA and DNA Is Dependent on Position in the Helix.” Biochemistry, 46(1), 172-181 (2007). Bevilacqua P. C.; Cerrone-Szakal A. L.; Siegfried N. A. “Insight Into the Functional Versatility of RNA Through Model-Making with Applications to Data Fitting.” Quarterly Reviews of Biophysics, 40, 55-85 (2007). Buckley T., Kitchen C., Siegfried N.A., Tefera E., Chen S., DiPaula B., Kelly D.L. “Novel Techniques for Therapeutic Drug Monitoring for Clozapine Levels.” Abstract for poster presentation, American Society of Health- Systems Pharmacists Midyear Clinical Meeting, December 2018, Anaheim, CA. Page 3 of 3

Honors and Awards

2011 – 2012 NIH F32, Ruth L. Kirschstein National Research Service Award 2009 – 2012 UNC Comprehensive Cancer Center Lineberger Fellowship 2008 Penn State Department of Chemistry Graduate Student Leadership and Service Award 2007 and 2008 Penn State Travel Award

2007 RNA Society Best Poster Award, Spring ACS National Meeting 2006 Penn State Apple Research Fellowship

2003 – 2004 Penn State Roberts Graduate Fellowship 2003 Moravian College Timothy M. Breidegam Memorial Student Service Award 2003 The American Institute of Chemists Award

1999 – 2003 Moravian College Comenius Scholar Fellowship 1999 – 2003 Moravian College JH Clewell Scholarship 2000 – 2003 PHEAA New Economy Technology Scholarship Skills and Qualifications

• Assay design, validation and troubleshooting • Effective team member and leader

• Method / technology transfer • Grant writing

• Project management • Writing of SOPs, organization of workflows

• Organized data collection and statistical analysis • Clear scientific writing and presentation

• Excellent communication skills • Directing collaborations with industry and academia

• Cross-functional team performance • Software proficiency: NCBI Tools (BLAST, genome browser), Adobe Photoshop & Illustrator, Excel, Word, PowerPoint, RNA Structure

Technical Expertise

Professional Memberships and Activities

Lineberger Comprehensive Cancer Center Postdoctoral Training Grant (NCI), Member: Postdoctoral Board o Organized a yearly one-day conference, including guest speaker and poster session o Oliver Smithies Nobel Symposium, Member: Organization Board o Wrote successful application for grant from the North Carolina Biotechnology Center RNA Society

American Association for Clinical Chemistry

The American Chemical Society (ACS)

Beta Beta Beta Biological Honor Society

Phi Eta Sigma National Academic Honor Society

Omicron Delta Kappa National Leadership Honor Society

• DNA / RNA extraction, purification,

manipulation

• Patient sample handling • UV/Vis and fluorescence spectroscopy

• GLP/GMP protocols, documentation • Digital droplet PCR, qPCR, rt-PCR, emulsion, touch-down, others

• Sample matrix reduction

• Immunoassays, ELISA, ImmunoPCR • RNA folding / manipulation / biochemistry • Large molecule structure determination

• RNA secondary structure analysis • Viral manipulation, growth assays • NGS library prep, sequencing analysis

• Sequencing (NGS and Sanger) • Fluorescence microscopy • NMR of RNA / DNA

• Tangential flow filtration

•

• Molecular diagnostics • Atomic absorption (AA) spectroscopy

• Mammalian cell and tissue culture • Qubit / Tape station / Bioanalyzer • DSC and ITC

• Clinical chemistry analyzers: Beckman

AU, Roche cobas series, Siemens Centaur

series

• Microparticle (magnetic or polystyrene)

handling, conjugation with proteins

• Chemical, biological, radioactive safety,

BSL 2 and 2 plus

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