A nanoscale plasmonic-gold platform for specific diagnosis of Zika and differentiation from other Flavivirus infections

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1 A nanoscale plasmonic-gold platform for specific diagnosis of Zika and differentiation from other Flavivirus infections Benjamin Pinsky, MD, PhD Assistant Professor of Pathology and Medicine Medical Director, Clinical Virology Laboratory Stanford University School of Medicine

2 Disclosures The assays described herein were developed by Nirmidas Biotech, Inc. in Palo Alto, CA and are currently available as research use only (RUO) reagents. Significant contributors to this project include Jesse Waggoner, Malaya Sahoo, and Janaki Abeynayake No personal financial interests to disclose. 2

3 Ancient Nanotechnology meets an Emerging Infectious Disease Gold (Au) nanoparticles have been used in art for centuries due to their brilliant color. Au nanoparticles have been used for decades in rapid antigen and antibody assays The Lycurgus Cup, 4 th Century - The British Museum Amendola et al. J Phys: Condens Matter

4 Gold Nanoparticles undergo Plasmonic Resonance The field of plasmonics represents the study of the interaction between light and conduction electrons of a metal. Plasmonic resonance is the collective oscillation of free electrons. Metal enhanced fluorescence can be used for analyte detection. Strobbia et al., Opt. Eng

5 Near Infrared Fluorescence Enhanced (NIR- FE) Imaging on Plasmonic Gold Tortuous, Elongated Au islands Cy5 IR800 Tabakman et al. Nature Comm

6 Near Infrared Fluorescence Enhanced (NIR- FE) Imaging on Plasmonic Gold continued x Tabakman et al. Nature Comm 2011 ZIKV IgG x ZIKV IgA Zhang and Pinsky et al. Nature Med

7 Nanoscale Plasmonic-Gold (pgold) Platform for Antibody Detection Zhang and Pinsky et al. Nature Med

8 ZIKV/DENV pgold Tests Antigens: ZIKV NS1 DENV-2 whole virus Up to 12 antigens per well Specimen Volume: <5 µl serum 16 wells per slide ~2 hour turnaround time Slides read on MidaScan-IR Dual channel (700 nm and 800 nm) near-infrared (NIR) confocal microscope scanner Images Courtesy of Meijie Tang, Nirmidas Biotech, Inc. 8

9 pgold IgM Assays Do Not Distinguish Between ZIKV and DENV Zhang and Pinsky et al. Nature Med

10 pgold ZIKV IgG distinguishes ZIKV from DENV infection Zhang and Pinsky et al. Nature Med

11 Zika virus: A new TORCH infection? ZIKV linked to the development of neonatal microcephaly and neurologic abnormalities ZIKV identified in amniotic fluid and fetal tissues. Johansson et al., NEJM 2016 Brasil et al., NEJM 2016 Rasmussen et al., NEJM 2016 Waggoner and Pinsky, J Clin Microbiol

12 Diagnosis of the TORCH Infections Toxoplasma Others (Syphilis, Parvovirus B19, HBV, HIV) Rubella Cytomegalovirus Herpes Simplex Virus Maternal Evaluation of the Timing of Exposure 12

13 IgG Avidity can be used to estimate the Timing of Exposure Avidity takes into account the presence of multiple Ag binding sites per Ab As B-cells undergo affinity maturation, antibody avidity increases Anticipate using ZIKV IgG + avidity to rule out exposure during pregnancy 13

14 pgold ZIKV IgG Avidity is consistent with recent infection Zhang and Pinsky et al., Nature Med

15 DENV pgold Avidity Detection reveals ZIKV infected patients were DENV experienced Zhang and Pinsky et al., Nature Med

16 Clinical Evaluation Needs and Challenges Nirmidas pgold ZIKV IgG and IgG Avidity Pregnancy Evaluate in pregnant women using longitudinal serum samples Capture the timing of seroconversion and increase in avidity Characterize the assay in DENV-experienced and naive populations. Characterize the ability to distinguish between other flavivirus and alphavirus antibodies. Multicenter, if possible, to account for host/virus variation Development of a 1 st trimester maternal screening panel IgG/IgG Avidity/ (IgM):ZIKV, Toxoplasma, CMV, (DENV) IgG: Rubella, T. pallidum, Varicella Antigen: HBV surface Ag, ZIKV Ag, (DENV Ag) 16

17 Clinical Evaluation Needs and Challenges Continued Other applications Test expanded numbers of adult and pediatric populations Confirm past infection (IgG/IgG Avidity) Potentially Identify early infection (IgG/IgA) Multiplex for antibody detection of other flaviviruses/alphaviruses ZIKV/CHIKV/DENV/YFV/MAYV Multiplex flavivirus/alphavirus antigen detection 17

18 Intended Use, Level of Care, and Target User Intended Use To rule out ZIKV exposure during pregnancy To identify recent infections Research/Epidemiologic Testing to determine seroprevalence Level of Care Clinical Laboratories (further automation required for near care testing) Public Health Laboratories Target User Institutional: Outpatient Primary Care, Travel, and Obstetrics Clinics Public Health/Research Epidemiology Individual: Clinical Laboratory Scientists 18

19 Summary Nirmidas pgold ZIKV/DENV Multiplexed assay on nanostructured plasmonic gold for the detection and distinction of IgG antibodies from patients infected with ZIKV and DENV. IgG Avidity may be useful to determine the timing of exposure, particularly in pregnancy. Further development requires evaluation of well characterized specimens 19

20 Acknowledgements Nirmidas Biotech, Inc. Bo Zhang Jey Ananata Meijie Tang Hongjie Dai Jesse Waggoner Funders: Stanford Department of Pathology National Institutes of Health Malaya Sahoo Janaki Abeynayake 20

21 Thank You! Dr. Waggoner Dr. Pinsky 21