Journal: Science Advances

Article Title: Virus detection using nanoparticles and deep neural network–enabled smartphone system

doi: 10.1126/sciadv.abd5354

Figure Lengend Snippet: Pt-nanoprobes preparation: ( A ) TEM image and particle size distribution histogram. ( B ) Schematic of Pt-nanoprobes structure and UV-vis absorption analysis showing spectra of PtNPs (citrate-capped PtNPs, black) and Pt-nanoprobes (PtNPs conjugated to mAb, red). Photo credit: Mohamed S. Draz, Brigham and Women’s Hospital. Virus capture and labeling on-chip: ( C ) SDS-PAGE analysis of ZIKV released from the surface of the chip. Lane M, protein marker; lane ZIKV on chip, ZIKV released from the chip; lane ZIKV, ZIKV standard solution (10 6 particles/ml, virus not added to the chip). Rectangles in red and blue edges mark bands from antibodies and ZIKV, respectively. ( D ) ELISA and RT-PCR assay results to detect and quantify the captured ZIKV on-chip. Error bars are SDs from two independent experiments. ( E ) SEM images of ZIKV captured on chip and labeled with Pt-nanoprobes: (i) ZIKV on chip and (ii) ZIKV captured and labeled with Pt-nanoprobes. The concentration of the virus sample was 10 6 particles/ml. The results are expressed as the average values of at least three independent experiments.

Article Snippet: Different sets of platinum nanoprobes were prepared of spherical PtNPs modified with mAb against the target virus: anti-ZIKV (ZIKV-Env) antibody (EastCoast Bio Inc. North Berwick, ME, USA) for ZIKV, anti-HBV surface antigen (Ad/Ay) antibody (HB24, catalog no. ab54247, Abcam, Cambridge, MA, USA) for HBV, and anti-HCV core antigen antibody (catalog no. ab2582, Abcam, Cambridge, MA, USA) for HCV.

Techniques: Virus, Labeling, SDS Page, Marker, Enzyme-linked Immunosorbent Assay, Reverse Transcription Polymerase Chain Reaction, Concentration Assay

Journal: Science Advances

Article Title: Virus detection using nanoparticles and deep neural network–enabled smartphone system

doi: 10.1126/sciadv.abd5354

Figure Lengend Snippet: ( A ) Digital images of the developed CNN-NES (i) and the actual microchips (ii) used to qualitatively test a virus-spiked sample. The system can measure the probability value of sample for qualitative testing (positive or negative) without the need for any optical smartphone attachment. Photo credit: Mohamed S. Draz, Brigham and Women’s Hospital. ( B ) ROC analysis of the developed CNN-NES compared to the standard RT-PCR using (i) ZIKV-spiked samples ( n = 60), (ii) HBV-spiked samples ( n = 22), (iii) ZIKV-infected patient serum samples ( n = 25), and (iv) HCV-infected patient serum samples ( n = 27). ( C ) Vertical scatterplots representing the CNN-NES probability values of the samples versus qualitative PCR results: (i) ZIKV-spiked samples ( n = 60), HBV-spiked samples ( n = 22), (ii) ZIKV-infected patient serum samples ( n = 25), and HCV-infected patient serum samples ( n = 27). ( D ) Vertical scatterplots representing the CNN-NES probability values of the samples versus quantitative PCR results: (i) ZIKV-spiked samples ( n = 60), HBV-spiked samples ( n = 22), (ii) ZIKV-infected patient serum samples ( n = 25), and HCV-infected patient serum samples ( n = 27).

Article Snippet: Different sets of platinum nanoprobes were prepared of spherical PtNPs modified with mAb against the target virus: anti-ZIKV (ZIKV-Env) antibody (EastCoast Bio Inc. North Berwick, ME, USA) for ZIKV, anti-HBV surface antigen (Ad/Ay) antibody (HB24, catalog no. ab54247, Abcam, Cambridge, MA, USA) for HBV, and anti-HCV core antigen antibody (catalog no. ab2582, Abcam, Cambridge, MA, USA) for HCV.

Techniques: Virus, Reverse Transcription Polymerase Chain Reaction, Infection, Real-time Polymerase Chain Reaction

Journal: Science Advances

Article Title: Virus detection using nanoparticles and deep neural network–enabled smartphone system

doi: 10.1126/sciadv.abd5354

Figure Lengend Snippet: ( A ) Custom-built microfluidic cartridge used for simple sample processing. ( B ) Exploded three-dimensional schematic of the developed cartridge indicating the detailed layer structure and the configuration of main components of the cartridge. ( C ) ROC curve analysis of CNN-NES enabled with the developed cartridge compared to the standard RT-PCR using PBS samples spiked with viruses, including HBV ( n = 42) and ZIKV ( n = 36). The samples with viral loads are classified as positive (≥250 particles/ml) and negative (<250 particles/ml). ( D ) Vertical scatterplot 1× PBS-spiked samples. The threshold for the probability values measured was 0.5. Samples with probability values above and below 0.5 were classified as positive (1) and negative (0), respectively, compared to the RT-PCR at a threshold of 250 particles/ml. ( E ) Evaluation of the repeatability of qualitative detection of samples when different cartridges were used for testing the same HBV-spiked PBS samples ( n = 25). The results are expressed as the average values of at least three measurements from one sample. ( F ) Evaluation of the performance of the system when operated by different users for sample testing using PBS samples spiked with ZIKV ( n = 18) and HBV ( n = 18). The results are expressed as the average values of three experiments from one sample. ( G ) Evaluation of the performance of the system when different smartphones were used for sample testing. The results are expressed as the average values of three independent experiments. The probability value is presented for each sample being positive. Error bars represent SDs. ns, not significant.

Article Snippet: Different sets of platinum nanoprobes were prepared of spherical PtNPs modified with mAb against the target virus: anti-ZIKV (ZIKV-Env) antibody (EastCoast Bio Inc. North Berwick, ME, USA) for ZIKV, anti-HBV surface antigen (Ad/Ay) antibody (HB24, catalog no. ab54247, Abcam, Cambridge, MA, USA) for HBV, and anti-HCV core antigen antibody (catalog no. ab2582, Abcam, Cambridge, MA, USA) for HCV.

Techniques: Reverse Transcription Polymerase Chain Reaction

Journal: Emerging Infectious Diseases

Article Title: Zika Virus RNA Replication and Persistence in Brain and Placental Tissue

doi: 10.3201/eid2303.161499

Figure Lengend Snippet: Oligonucleotide primers used for RT-PCR assays*

Article Snippet: Zika virus ISH was performed by using sense and antisense riboprobes that target multiple genes of Zika virus (Advanced Cell Diagnostics, Newark, CA, USA).

Techniques: Sequencing, Virus

Journal: Emerging Infectious Diseases

Article Title: Zika Virus RNA Replication and Persistence in Brain and Placental Tissue

doi: 10.3201/eid2303.161499

Figure Lengend Snippet: Symptom onset trimester, pregnancy outcomes and Zika virus tissue RT-PCR results of 52 case-patients*

Article Snippet: Zika virus ISH was performed by using sense and antisense riboprobes that target multiple genes of Zika virus (Advanced Cell Diagnostics, Newark, CA, USA).

Techniques: Virus

Journal: Emerging Infectious Diseases

Article Title: Zika Virus RNA Replication and Persistence in Brain and Placental Tissue

doi: 10.3201/eid2303.161499

Figure Lengend Snippet: Characteristics and laboratory findings for 13 microcephaly-associated case-patients*

Article Snippet: Zika virus ISH was performed by using sense and antisense riboprobes that target multiple genes of Zika virus (Advanced Cell Diagnostics, Newark, CA, USA).

Techniques: Virus, Membrane

Journal: Emerging Infectious Diseases

Article Title: Zika Virus RNA Replication and Persistence in Brain and Placental Tissue

doi: 10.3201/eid2303.161499

Figure Lengend Snippet: Characteristics and laboratory findings of case-patients with adverse pregnancy outcome*

Article Snippet: Zika virus ISH was performed by using sense and antisense riboprobes that target multiple genes of Zika virus (Advanced Cell Diagnostics, Newark, CA, USA).

Techniques: Virus, Membrane

Journal: Emerging Infectious Diseases

Article Title: Zika Virus RNA Replication and Persistence in Brain and Placental Tissue

doi: 10.3201/eid2303.161499

Figure Lengend Snippet: Zika virus RNA load levels in human brain and placental tissues. The scatter plot graph shows the relative levels of Zika virus RNA in formalin-fixed, paraffin-embedded tissue sections, which were quantified by real-time quantitative reverse transcription PCR by using primer-probe sets for Zika virus envelope gene and β-actin mRNA. β-actin mRNA was used as an internal reference gene that provided a normalization factor for the amount of RNA extracted from a section. The copy number of Zika virus RNA per cell was calculated using β-actin mRNA copy number, which was estimated to be 1,500 copies/cell. The graph shows individual data points and superimposed horizontal lines at the geometric mean, and error bars show the 95% CI for that geometric mean. p values were calculated with nonparametric 1-way analysis of variance (Kruskal-Wallis test) followed by Dunn multiple comparison tests. The relative Zika virus RNA copy numbers for second/third trimester or full-term placentas were statistically significantly lower than those for first trimester placentas or infant brain tissues.

Article Snippet: Zika virus ISH was performed by using sense and antisense riboprobes that target multiple genes of Zika virus (Advanced Cell Diagnostics, Newark, CA, USA).

Techniques: Virus, Formalin-fixed Paraffin-Embedded, Reverse Transcription, Comparison

Journal: Emerging Infectious Diseases

Article Title: Zika Virus RNA Replication and Persistence in Brain and Placental Tissue

doi: 10.3201/eid2303.161499

Figure Lengend Snippet: Localization of Zika virus RNA by in situ hybridization in brain tissues from infants with microcephaly. A) ISH with use of antisense probe. Zika virus genomic RNA (red stain) in cerebral cortex of an infant (case-patient no. 66, gestational age 26 wk). Original magnification ×10. B) ISH with use of sense probe. Serial section showing negative-strand replicative RNA intermediates (red stain) in the same areas shown in panel A. Original magnification ×10. C) ISH with use of antisense probe. Higher magnification of panel A, showing cytoplasmic staining of neural (arrowheads) and glial cells. Original magnification ×20. D) ISH with use of sense probe. Higher magnification of panel B, showing cytoplasmic staining of neural and glial cells (arrowheads). Original magnification ×20. E) ISH with use of antisense probe. Localization of negative-strand replicative RNA intermediates in neural cells or neurons (red, arrowheads) of another infant with fatal outcome (case-patient no. 67, gestational age 27 wk). Original magnification ×40. F) Immunostaining of neurons (arrowheads) with use of antibodies against neuronal nuclei in a serial section. Original magnification ×40. G) Hematoxylin and eosin stain showing cortical neural cells in a serial section. Original magnification ×40. H) Immunostaining of glial cells (arrowheads) with use of glial fibrillary acidic protein antibody in the same case. Original magnification ×40. ISH, in situ hybridization.

Article Snippet: Zika virus ISH was performed by using sense and antisense riboprobes that target multiple genes of Zika virus (Advanced Cell Diagnostics, Newark, CA, USA).

Techniques: Virus, RNA In Situ Hybridization, Staining, Immunostaining, H&E Stain, In Situ Hybridization

Journal: Emerging Infectious Diseases

Article Title: Zika Virus RNA Replication and Persistence in Brain and Placental Tissue

doi: 10.3201/eid2303.161499

Figure Lengend Snippet: Localization of Zika virus RNA by ISH in placental tissues of women after spontaneous abortion. A) ISH with use of antisense probe. Zika virus genomic RNA localization in placental chorionic villi, predominantly within Hofbauer cells (red stain, arrows), of a case-patient who had spontaneous abortion at 11 wk gestation (case-patient no. 56). Original magnification ×10). B) ISH with use of sense probe. Serial section showing negative–strand replicative RNA intermediates (red stain, arrows) in the same cells shown in panel A. Original magnification ×10. C) Hematoxylin and eosin stain of placental tissue of a case-patient who experienced spontaneous abortion at 8 wk gestation (case-patient no. 47). Original magnification ×20. D) Immunostaining for CD163 highlighting villous Hofbauer cells in a serial section as seen in panel C. Original magnification ×63. E) ISH with use of antisense probe. Zika virus genomic RNA as seen in a serial section from the same case-patient as in panel C, showing staining within Hofbauer cells (red stain, arrows) of placental chorionic villi. Original magnification ×40. F) ISH with use of sense probe. Serial section showing negative-strand replicative RNA intermediates (red stain, arrows) in the same cells as shown in panel E. Original magnification ×40. G) Hematoxylin and eosin stain from the same case-patient as in panel C, showing inflammatory cell infiltrates in maternal side of placenta. Original magnification ×63. H) ISH with use of sense probe. Negative-strand replicative RNA intermediates (red stain, arrows) in inflammatory cells in a serial section. Original magnification ×63. ISH, in situ hybridization.

Article Snippet: Zika virus ISH was performed by using sense and antisense riboprobes that target multiple genes of Zika virus (Advanced Cell Diagnostics, Newark, CA, USA).

Techniques: Virus, Staining, H&E Stain, Immunostaining, In Situ Hybridization