Supplementary Materials for
Zika virus activates de novo and cross-reactive memory B cell responses
in dengue-experienced donors
Thomas F. Rogers, Eileen C. Goodwin, Bryan Briney, Devin Sok, Nathan Beutler,
Alexander Strubel, Rebecca Nedellec, Khoa Le, Michael E. Brown,
Dennis R. Burton,* Laura M. Walker*
*Corresponding author. Email: [email protected] (L.M.W.); [email protected] (D.R.B.)
Published 18 August 2017, Sci. Immunol. 2, eaan6809 (2017)
DOI: 10.1126/sciimmunol.aan6809
This PDF file includes:
Fig. S1. Binding of acute serum to ZIKV and DENV1–4 viral particles and NS1
proteins.
Fig. S2. Plasmablast sorting.
Fig. S3. Binding of plasmablast-derived mAbs to ZIKV-Para by ELISA.
Fig. S4. Western blot analysis of mAb binding to ZIKV lysate.
Fig. S5. Heat map of mAb binding reactivity to recombinant ZIKV E, WNV E,
DIII, and E-FL.
Fig. S6. mAb binding to recombinant ZIKV and DENV E proteins.
Fig. S7. ELISA binding of whole virus–specific clones to ZIKV and DENV.
Fig. S8. SHM of NS1-specific mAbs.
Fig. S9. Representative neutralization curves.
Fig. S10. Enhancement of ZIKV infection by plasmablast-derived mAbs isolated
from DENV-experienced donors.
Fig. S11. Memory B cell sorting.
Fig. S12. Distribution of clonal lineage members across plasmablast and memory
B cell subsets.
Fig. S13. Epitope mapping of ZIKV-specific mAbs derived from memory B cells.
Table S1. Clinical characteristics of ZIKV-infected donors.
Table S2. Binding properties of ZIKV E–specific mAbs isolated from
plasmablasts.
Table S3. Binding characteristics of mAb 4G2.
Table S4. Sequences of ZV-67 competitor mAbs.
immunology.sciencemag.org/cgi/content/full/2/14/eaan6809/DC1
Table S5. Competition of ADI-24247 and ADI-24314 with each other and with
control mAbs.
Table S6. Binding properties of NS1-specific mAbs isolated from plasmablasts.
Table S7. Neutralizing activity of selected plasmablast-derived mAbs.
Table S8. Binding properties of ZIKV E–specific mAbs isolated from memory B
cells.
Table S9. Clonal lineages shared between plasmablast and memory B cell–
derived antibodies.
Table S10. Neutralizing activity of selected memory B cell–derived mAbs.
Fig. S1. Binding of acute serum to ZIKV and DENV1 4 viral particles and NS1 proteins. Acute serum from ZIKV-infected donors were tested by ELISA for the presence of IgG antibodies to A) whole ZIKV and DENV1-4 particles and B) recombinant ZIKV and DENV1-4 NS1 proteins. C) Acute serum neutralization of ZIKV-Para and DENV1-4. Neutralization assays were performed using a live virus plaque reduction assay. IC50 values represent the serum dilution at which viral infectivity is reduced by 50%.
10-5 10-4 10-3 10-2 10-1 1000
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OD4
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ZIKV-Para
DENV3
DENV1DENV2
DENV4
Donor 1 Donor 2 Donor 3 Donor 4
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A)
B)
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C)
D) Serum Neutralization IC50 (1/dilution)
ZIKV-ParaDonor DENV-1 DENV-2 DENV-3 DENV-4901 <10 10 21 10
2 212 84 59 84 1803 1350 3078 87714343 37264 925 3465 695 400 1120
–
Fig. S2. Plasmablast sorting. Representative fluorescence activated cell sorting (FACS) gating strategy used for plasmablast sorting. Lymphocytes were gated based on forward and side scatter, followed by a live/dead gate and selection of CD3− CD8− CD14− cells. B cells were identified by gating on CD19+ and CD20+ cells. Plasmablasts, defined as CD19+ CD3− CD20−/lo CD38hi CD27hi cells, were single-cell sorted for mAb cloning. SSC-A, side scatter area; FSC-A, forward scatter area.
Fig. S3. Binding of plasmablast-derived mAbs to ZIKV-Para by ELISA. Purified IgGs from donors 1, 2, 3, and 4 were tested for binding to live ZIKV-Para at a concentration of 10 µg/ml. Antibodies with ELISA OD405 values greater than 0.3 (dotted line) were considered binders. MAb 4G2 was included as a positive control and is shown as a red bar.
1 5 9 13 17 21 25 29 33 37 41 45 49 53 57 61 65 69 73 77 81 85 89 93 9710110
510
911
311
70
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Donor 1
1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 51 53 55 57 59 610
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1 2 3 4 5 6 7 8 91011121314151617181920212223242526272829303132333435360
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1 3 5 7 9 111315171921232527293133353739414345474951535557596163656769710
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OD4
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Donor 3
Donor 2
Donor 4
Fig. S4. Western lot analysis of mAb binding to ZIKV lysate. Western Blots performed under reducing conditions show reactivity of plasmablast-derived mAbs with ZIKV E and prM proteins derived from whole zika virus lysate. 4G2 was included as a positive control for reactivity to recombinant (lane #1) and whole virus-derived (lane #2) E protein. A prM-specific mAb (GTX128092) was included as a positive control for reactivity to prM from DENV-1 whole virus lysate (lane #3).
15
2025
37
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150kDa
Env ZIKV DENV-1 ZIKV
4G2
4G2
Anti-prM
ADI-2
4234
ADI-2
4223
ADI-2
4225
ADI-2
4250
ADI-2
4251
ADI-2
4286
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4334
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ADI-2
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ADI-2
4209
ADI-2
4273
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2025
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150kDa
Env ZIKV DENV-1 ZIKV
4G2
4G2
Anti-prM
ADI-2
4209
ADI-2
4235
ADI-2
4240
ADI-2
4253
ADI-2
4268
ADI-2
4270
ADI-2
4271
ADI-2
4273
ADI-2
4274
ADI-2
4222
ADI-2
4289
ADI-2
4294
ADI-2
4295
ADI-2
4302
ADI-2
4318
ADI-2
4320
ADI-2
4323
ADI-2
4329
ADI-2
4341
ADI-2
4351
ADI-2
4288
ADI-2
4369
ADI-2
4371
ADI-2
4372
ADI-2
4374
b
Fig. S5. Heat map of mAb binding reactivity to recombinant ZIKV E, WNV E, DIII, and E-FL. Plasmablast-derived mAbs from three DENV-experienced donors were testing for binding to recombinant ZIKV E, WNV E, DIII and E-FL by BLI. Apparent mAb binding EC50s are shown in the heat map.
ZIKV EE-FLWNV EDIII
0.1-1 nM
1-10 nM
10-100 nM
100-500 nM
>500 nM
Fig. S6. mAb binding to recombinant ZIKV and DENV E proteins. Plasmablast-derived mAbs from three DENV-experienced donors were tested for binding to recombinant ZIKV and DENV1-4 E proteins. Apparent binding affinities were determined by BLI measurements. Each point represents a single mAb. Statistical comparisons were made using the Mann-Whitney test (***P < 0.001) and Kruskal-Wallis test (P <0.001).
0.1
1
10
100
1000
IgG
KD
(nM
)
******
******
ZIKV E
DENV-1 E
DENV-2 E
DENV-3 E
DENV-4 E
Fig. S7. ELISA binding of whole virus-specific clones to ZIKV and DENV. A) A subset of whole virus-specific mAbs derived from three DENV-experienced donors were titrated against live ZIKV and DENV particles. B) Whole-virus specific mAbs from a ZIKV infected DENV-naïve donor were tested for binding to whole ZIKV-Para and DENV1-4 particles at a concentration of 10 µg/ml. The heat map shows the ELISA binding OD at this concentration. Antibodies with ELISA OD405 values greater than 0.3 were considered binders. The pan-flavivirus mAb 4G2 was included as a positive control.
0.0001 0.001 0.01 0.1 1 100
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ADI-24323
IgG concentration (ug/ml)
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ADI-24318
IgG concentration (ug/ml)
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ADI-24329
IgG concentration (ug/ml)
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ADI-24251
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ADI-24320
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ZIKV AmericaDENV1DENV2DENV3DENV4
A)
B)
ZIKV
-Par
aDE
NV1
DENV
2DE
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DENV
4
ELISA OD at 10 ug/ml
Donor 1 mAbs
4G2 control
0 - 0.3
0.3 - 0.5
0.5 - 1.0
1.0 - 4.0
Fig. S8. S of NS1-specific mAbs. Load of somatic mutations (expressed as number of nucleotide substitutions in the variable region of the heavy chain) in ZIKV NS1-specific mAbs isolated from plasmablasts. Each point represents an individual mAb. Red bars indicate the median number of nucleotide substitutions.
Donor 1
Donor 2
Donor 3
Donor 4
0
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40
50
VHnucleotidesubstitutions
(#)
HM
Fig. S9. Representative neutralization curves. Neutralization titration experiments were performed using a live virus plaque reduction assay.
10 -6 10 -4 10 -2 10 00
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ZIKV-Para
241922423224257
2437024361
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2436124326
24366
Fig. S10. Enhancement of ZIKV infection by plasmablast-derived mAbs isolated from DENV-experienced donors. (a) Correlation between neutralization potency and concentration of mAb at peak enhancement, defined as the concentration of mAb that resulted in the highest number of ZIKV foci when K562 cells were infected. (b) Correlation between epitope specificity and concentration of mAb at peak enhancement with mean and standard deviation displayed as line and bars, respectively. (c) Correlation between neutralization potency and the sum of enhancement activity over the range of antibody concentrations tested as defined by area under the curve (AUC). Enhancement assays were performed using non-permissive K562 cells. Statistical comparisons were made using Spearman correlation and Kruskal-Wallis test.
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[mAb] at peak enhancement of ZIKV-Para
ZIKV
-Par
a IC
50p = 0.0001
DIII FL
Whole
virus
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Unkno
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] at p
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IKV-
Para
A) B)
C)
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AUC of enhancement of ZIKV-Para
ZIKV
-Par
a IC
50
p = 0.0096
Fig. S11. Memory B cell sorting. Representative fluorescence activated cell sorting (FACS) gating strategy used for ZIKV E-specific memory B cell sorting. Lymphocytes were gated based on forward and side scatter, followed by a live/dead gate and selection of CD3− CD8− CD14− cells. B cells were identified by gating on CD19/CD20+ cells. IgG+ B cells that showed reactivity with ZIKV E (right-most plot, quadrant 2) were single-cell sorted for mAb cloning. SSC-A, side scatter area; FSC-A, forward scatter area.
FSC-A
SSC
-A
Dum
p
CD
27-B
V421
ZIKV
A-A
PC
CD19/CD20-PeCy7 IgG-BV605 ZIKV E-PE
13.0
95.255.2
0.86
0.05299.0
0.037
Fig. S12. Distribution of clonal lineage members across plasmablast and memory B cell subsets. Ribbons linking arcs represent shared clonal lineages. The thickness of each ribbon is proportional to the number of clones in the lineage. Circular visualization plots were created using circos v0.64 (http://circos.ca/). Donor 3 lineages are shown in the plot. No clonal lineages were shared across plasmablast and memory B cell-derived responses from donor 4.
Memory B cellPlasmablast
Fig. S13. Epitope mapping of ZIKV-specific mAbs derived from memory B cells. Pie charts show the distribution of mAbs that bind to epitopes within DIII, epitopes within or proximal to the fusion loop on DII, epitopes defined by ADI-30056, ADI-30023, or ADI-24314, and undefined epitopes. Antibody competition assays were used to define epitopes.
22
Donor 4UnknownADI-24314FLDIII
51
Donor 3
ADI-30056ADI-30023
Table S1. Clinical characteristics of ZIKV-infected donors.
Donor Date of symptom onset Age Gender ZIKV PCR Country of origin (Region of origin)
Country of exposure (Region of exposure)
First collection date Second collection date
1 12/29/2015 55 M + United States (Texas) Aruba 1/8/2016 n/a2 2/18/2016 46 M + Columbia (Barranquilla) Columbia (Barranquilla) 2/26/2016 7/28/20163 2/19/2016 51 F + Columbia (Barranquilla) Columbia (Barranquilla) 2/26/2016 7/28/20164 2/20/2016 30 F + Columbia (Barranquilla) Columbia (Barranquilla) 2/26/2016 7/28/2016
Table S2. Binding properties of ZIKV E-specific mAbs isolated from plasmablasts.
Clone Donor DIII E-FL WNV E ZIKV E DENV-1 E DENV-2 E DENV-3 E DENV-4 E ZV67 4G2 ADI-24314 ADI-24247 Epitope designation
ADI-32211 1 ND ND ND 3.29E-08 N.B. N.B. N.B. N.B. ND ND ND ND NC
ADI-24249 2 N.B. 1.4E-08 N.B. 3.43E-08 3.82E-09 1.86E-09 4.88E-06 2.03E-09 0.96 1.46 2.08 5.3 ADI-24247
ADI-24247 2 N.B. 1.1E-09 1.14E-09 1.45E-09 2.95E-09 1.04E-09 1.85E-09 5.90E-10 1.01 1.40 1.48 53.2 ADI-24247
ADI-24230 2 N.B. 4.2E-09 N.B. 1.69E-08 2.51E-09 2.06E-09 3.75E-06 2.76E-09 1.01 1.30 2.17 5.9 ADI-24247
ADI-24234 2 N.B. 6.2E-10 1.36E-09 2.64E-09 1.38E-09 1.07E-09 7.43E-10 1.11E-09 1.07 1.31 1.41 70.7 ADI-24247
ADI-24246 2 N.B. 4.5E-10 9.12E-10 2.44E-09 2.25E-09 1.25E-09 8.02E-10 1.49E-09 1.09 1.36 1.27 24.4 ADI-24247
ADI-24244 2 N.B. 5.4E-10 6.43E-10 2.74E-09 2.95E-09 6.14E-10 5.83E-10 4.94E-10 1.14 1.45 1.41 77.7 ADI-24247
ADI-24245 2 N.B. 1.5E-09 3.26E-09 3.47E-09 3.12E-09 1.01E-09 2.20E-09 5.07E-10 1.17 1.43 1.44 40.7 ADI-24247
ADI-24202 2 N.B. 9.6E-10 1.71E-09 6.04E-09 3.83E-09 1.22E-09 1.30E-09 1.07E-09 1.39 1.48 2.20 36.4 ADI-24247
ADI-24200 2 N.B. 4.2E-09 N.B. 7.19E-09 3.04E-09 N.B. 1.82E-08 6.41E-09 1.27 1.60 34.56 1.5 ADI-24314
ADI-24194 2 N.B. 1.4E-08 5.40E-08 2.07E-08 1.66E-08 3.42E-08 2.92E-09 4.21E-09 1.30 2.72 50.49 1.5 ADI-24314
ADI-24229 2 1.85E-08 6.9E-10 N.B. 2.39E-08 6.73E-10 N.B. N.B. N.B. 2.2 2.7 1.8 1.3 DIII
ADI-24197 2 1.418E-08 W.B. N.B. N.B. 2.08E-09 N.B. N.B. N.B. N/A N/A N/A N/A DIII
ADI-24232 2 6.84697E-09 6.7E-10 N.B. 4.49E-08 8.84E-10 N.B. N.B. N.B. 4.4 1.8 N/A N/A DIII (ZV-67 competitor)
ADI-24191 2 5.233E-09 1.2E-08 N.B. 4.51E-08 1.14E-08 N.B. N.B. N.B. 6.0 3.2 N/A N/A DIII (ZV-67 competitor)
ADI-24227 2 2.155E-09 3.4E-09 N.B. 5.93E-08 1.54E-08 N.B. N.B. N.B. 6.5 1.4 N/A N/A DIII (ZV-67 competitor)
ADI-24192 2 4.086E-09 8.6E-09 N.B. 5.28E-08 7.92E-10 N.B. N.B. N.B. 7.5 1.5 N/A N/A DIII (ZV-67 competitor)
ADI-24238 2 1.4771E-09 1.2E-09 N.B. 2.39E-08 4.67E-09 N.B. N.B. N.B. 10.3 1.3 N/A N/A DIII (ZV-67 competitor)
ADI-24255 2 1.12486E-09 5.0E-10 N.B. 2.67E-08 1.81E-09 N.B. N.B. N.B. 15.0 1.4 N/A N/A DIII (ZV-67 competitor)
ADI-24211 2 N.B. N.B. 4.49E-10 1.48E-09 1.40E-09 6.63E-10 4.62E-10 5.02E-10 0.80 66.76 N/A N/A FL (4G2-like)
ADI-24220 2 N.B. N.B. 3.51E-10 1.38E-09 8.72E-10 6.72E-10 4.39E-10 4.76E-10 0.87 103.18 N/A N/A FL (4G2-like)
ADI-24224 2 N.B. N.B. 2.58E-09 1.39E-09 1.25E-09 1.12E-09 6.56E-10 9.53E-10 0.90 112.69 N/A N/A FL (4G2-like)
ADI-24219 2 N.B. N.B. 3.44E-10 1.53E-09 1.06E-09 6.81E-10 4.79E-10 5.36E-10 0.96 127.06 N/A N/A FL (4G2-like)
ADI-24195 2 N.B. N.B. 5.61E-10 2.05E-09 2.35E-09 9.30E-10 6.52E-10 6.70E-10 0.98 73.34 N/A N/A FL (4G2-like)
ADI-24190 2 N.B. 8.3E-09 N.B. 8.15E-09 1.54E-09 1.31E-09 5.19E-10 4.87E-10 1.06 39.90 N/A N/A FL proximal
ADI-24237 2 N.B. N.B. N.B. 2.59E-07 N.B. N.B. N.B. N.B. N/A N/A N/A 1.3 NC
ADI-24231 2 N.B. N.B. N.B. N.B. N.B. N.B. N.B. 4.93E-08 N/A N/A N/A N/A NC
ADI-24198 2 N.B. 2.1E-09 N.B. 7.58E-09 1.28E-09 3.23E-09 1.31E-09 8.37E-10 1.10 1.76 2.36 1.6 NC
ADI-24254 2 N.B. N.B. N.B. N.B. N.B. N.B. N.B. 1.22E-07 N/A N/A N/A 3.0 NC
ADI-24269 3 N.B. 1.6E-09 1.48E-07 2.17E-09 1.25E-09 4.08E-10 5.29E-10 2.96E-10 1.07 1.90 79.19 1.8 ADI-24314
ADI-24276 3 N.B. N.B. 4.69E-10 1.56E-09 8.76E-10 7.13E-10 5.39E-10 5.08E-10 0.84 80.40 N/A N/A FL (4G2-like)
ADI-24283 3 N.B. N.B. N.B. 4.18E-07 N.B. N.B. 1.08E-07 N.B. N/A N/A N/A 2.1 NC
ADI-24272 3 N.B. N.B. N.B. 5.88E-08 6.61E-09 5.86E-09 5.21E-09 4.56E-09 0.97 5.20 N/A N/A Non WNV FL
ADI-24257 3 N.B. 1.5E-08 N.B. 1.73E-08 N.B. N.B. N.B. N.B. 19.1 1.3 N/A N/A ZV-67 competitor
ADI-24285 3 N.B. N.B. 8.06E-10 1.30E-09 1.26E-09 1.29E-09 2.12E-09 1.92E-09 0.87 33.86 N/A N/A FL (4G2-like)
ADI-24296 3 N.B. N.B. 3.76E-10 1.25E-09 7.71E-10 5.95E-10 4.19E-10 4.82E-10 0.88 96.71 N/A N/A FL (4G2-like)
ADI-24292 3 N.B. N.B. N.B. 1.51E-07 3.02E-07 N.B. 4.38E-08 N.B. N/A N/A N/A 1.6 NC
ADI-24345 4 N.B. 1.4E-08 1.49E-08 3.14E-08 2.02E-09 9.24E-10 7.49E-10 5.61E-10 0.91 1.18 41.18 6.5 ADI-24314
ADI-24333 4 N.B. 7.5E-10 N.B. 2.10E-09 8.47E-10 5.91E-10 5.50E-10 1.39E-09 0.96 1.92 53.88 1.2 ADI-24314
ADI-24314 4 N.B. 1.3E-09 W.B. 3.57E-09 1.69E-09 9.30E-10 1.18E-09 1.13E-09 1.04 2.27 61.85 1.4 ADI-24314
ADI-24304 4 N.B. 2.2E-09 4.47E-09 5.91E-09 9.58E-10 1.22E-09 5.70E-10 6.15E-10 1.18 2.86 38.87 2.0 ADI-24314
ADI-24336 4 N.B. N.B. 6.41E-10 1.78E-09 7.52E-10 7.91E-10 9.71E-10 1.43E-09 0.90 59.63 N/A N/A FL (4G2-like)
ADI-24346 4 N.B. N.B. 2.60E-10 1.01E-09 7.68E-10 1.04E-09 2.05E-10 2.00E-10 0.94 102.44 N/A N/A FL (4G2-like)
ADI-24340 4 N.B. N.B. 4.62E-10 1.85E-09 2.05E-09 7.36E-10 1.13E-09 8.08E-10 0.95 99.36 N/A N/A FL (4G2-like)
ADI-24348 4 N.B. N.B. 6.50E-10 2.45E-09 1.22E-09 1.06E-09 6.92E-10 7.61E-10 0.97 76.21 N/A N/A FL (4G2-like)
ADI-24353 4 N.B. N.B. 6.83E-10 2.31E-09 9.82E-10 1.17E-09 7.79E-10 8.38E-10 1.02 65.89 N/A N/A FL (4G2-like)
ADI-24313 4 N.B. N.B. 2.85E-09 5.87E-09 3.02E-09 1.19E-08 1.79E-09 7.03E-09 1.04 24.16 N/A N/A FL (4G2-like)
ADI-24356 4 N.B. 2.8E-08 N.B. 6.01E-08 2.67E-09 N.B. N.B. N.B. 2.6 5.5 N/A N/A FL proximal
ADI-24370 4 N.B. N.B. N.B. W.B. N.B. 3.59E-08 N.B. N.B. 1.14 15.54 N/A N/A NC
ADI-24366 4 N.B. N.B. N.B. 1.38E-08 N.B. N.B. N.B. N.B. N/A N/A N/A N/A NC
ADI-24361 4 N.B. N.B. N.B. 1.47E-08 N.B. N.B. 5.65E-09 N.B. N/A N/A N/A 1.5 NC
ADI-24368 4 N.B. N.B. N.B. 2.08E-08 N.B. N.B. N.B. N.B. N/A N/A N/A N/A NC
ADI-24326 4 N.B. N.B. N.B. 2.24E-08 N.B. N.B. N.B. N.B. N/A N/A N/A N/A NC
ADI-24375 4 N.B. N.B. N.B. 2.82E-08 N.B. N.B. N.B. N.B. N/A N/A N/A 1.2 NC
ADI-24373 4 N.B. N.B. N.B. N.B. N.B. 5.57E-08 N.B. N.B. N/A N/A N/A N/A NC
ADI-24365 4 N.B. N.B. N.B. W.B. N.B. N.B. N.B. N.B. N/A N/A N/A N/A NC
ADI-24342 4 N.B. W.B. N.B. 8.25E-08 N.B. N.B. N.B. N.B. 0.99 1.61 1.31 1.4 NC
ADI-24343 4 N.B. N.B. N.B. 1.41E-07 N.B. N.B. N.B. N.B. 1.00 18.84 N/A N/A Non WNV FL
ADI-24352 4 N.B. N.B. N.B. 2.39E-09 1.00E-09 9.18E-10 7.77E-10 6.07E-10 1.08 46.02 N/A N/A Non WNV FL
ADI-24367 4 N.B. 2.1E-08 N.B. 4.99E-08 1.46E-09 N.B. N.B. N.B. 13.2 1.5 N/A N/A ZV-67 competitor
Apparent IgG KD (M)*Antibody competition (Fold reduction in antigen binding
in the presence of competitor Fab)#
* N.B., non-binder; W.B., weak binder. IgG KDs were calculated for MAbs with ForteBio binding responses >0.1 nm. MAbs with ForteBio binding responses <0.05 nm were designated as N.B.; MAbs with ForteBio binding responses between 0.05 and 0.1 nm were designated as W.B. # Results are expressed as the fold reduction in antigen binding in the presence of saturating concentrations of competitor Fab relative to an antigen-only control.
Table S3. Binding characteristics of mAb 4G2.
Antigen Rmax (RU) ka (1/Ms) kd (1/s) KD (pM)
ZIKV E 23.1 4.04E+05 1.62E-05 39.6ZIKV E-FL - - - N.B.
WNV E 19.4 1.17E+06 1.19E-04 102N.B., non-binder.
Table S4. Sequences of ZV-67 competitor mAbs.
Clone VH Germline VH FR1 VH CDR1 VH FR2 VH CDR2 VH FR3 VH CDR3
ADI-24191 VH3-23 QVTLKESGGGLVQPGGSRRLSCVASG FTFSSFAMS WVRQAPGKGLEWVS SITVDSGSKYYAGSVEG RFTISRANSRNTLYLQMNDLTAEDTAVYYC AKDRPLHGFGELYDHADI-24227 VH3-23 EVQLLESGGGLIQPGGSLRLSCAASG FTFSTYAMN WVRQAPGKGLEWVS SITASSDSTYYADSVKG RFTISRDNSRNTLYLQMNSLRAEDTALYYC AKDRPPYGVGELYDYADI-24238 VH3-23 EVQLLESGGGLVQPGGSVRLSCVASG FTFSKFAMS WVRQAPGKGLEWVS TIAASDDSTYYVDSVKG RFTISRANSKNTLYLQMNSLRVEDTAVYYC AKDRPPYGVGELYDYADI-24192 VH3-23 EVQLLESGGGLVQPGGSLRLSCAASG FPFRDYGMS WVRQAPGKGLEWVS AISSIDDSTYYAASVRG RFTISRDNSKNTLDLQMNSLRAEDTAVYFC AKDRSTRGFGELLNYADI-24229 VH3-23 EVQLVESGGGLVQPGGSLRLSCAASG FTFSSYGMS WVRQAPGKGLEWVS SLTGSGVDTYYAGSVKG RFTISRDNSKNTLYLQMNSLRVEDTAVYYC ARDRLGQGFGELFAFADI-24232 VH3-23 EVQLLESGGGLVQPGGSLRLSCAASG FTFSSYGMS WVRQAPGKGLQWVS AISSIDDSTYYADSVKG RFTISRDNSKNTLYLQMNSLRVADTALYYC GKDRTARGFGELLDSADI-24255 VH3-23 EVQLVESGGGLVQPGGSLRLSCVASG FSFSSYGLS WVRQAPGKGLQWVS GISSIDDSKYYADSVKG RFTISRDNSKNTLYLQMNSLTVADTALYYC GKDRVARGFGELLDSADI-24197 VH3-30 QVQLVESRGGVVQPGGSLRLSCVAPG FTFRVYDMH WVRQAPGKGLEWVA RIQYDGTNTYYADSVKG RLTISRDNSKNTLYLQMNSLRLEDTAVYYC ARDGYNHHLGYHHYMDV
Clone VL Germline VL FR1 VL CDR1 VL FR2 VL CDR2 VL FR3 VL CDR3
ADI-24191 VK1-27 DIQVTQSPSSLSASVGDRVTVTC RASQGISNYLA WYQQKPGKAPKLLIS GASTLQS GVPSRFSGSGSGTDFTLTISSLQPEDVATYYC QKYNSVPWTADI-24227 VK1-27 DIRVTQSPSSLSASVGDRVTITC RASQGISNYLA WYQQKPGKVPKLLIS AASTLQS GVPSRFSGSGSGTDFTLTISSLQPEDVATYYC QKYNSVPWTADI-24238 VK1-27 DIQMTQSPSSLSASVGDRVTITC RASQGISNYLA WYQQKPGKVPNLLIS AASTLQS GVPSRFSGSGSGTLFTLTISNLQPEDVATYYC QKYNSVPWTADI-24192 VK1-5 DIRVTQSPSTLSASVGDRVTITC RASQNINSWLA WYQQKPGKAPNLLIY KASSLES GVPLRFSGSGSGTEFTLTISRLQPDDFATYYC QHYYSYPWTADI-24229 VK1-5 DIQMTQSPSTLSASVGDRVTITC RASQSLSSWLA WYQQKPGKAPRLLIH KASILQS GVPLRFSGSGSGTEFTLTISSLQPDDFATYYC QHYYSYPWTADI-24232 VK1-5 DIRMTQSPSTLSASVGDRVTITC RASQSISSWLA WFQQKPGKAPKLLIF KASSLQN GVPSRFSGSGSGTEFTLTISSLQPDDFATYYC QHYYSYPWTADI-24255 VK1-5 DIRLTQSPSTLSASVGDRVTITC RASQSINSWLA WFQQKPGKAPNLLMF KASTLHT GVPSRFSGSGSGTEFTLTISSLQPDDFATYYC QHYYSYPWTADI-24197 VK3-20 EIVLTQSPGTLSLSPGERATLSC RASQSVSSTSLV WYQQKPGQPPRLLIY SASRRTS GVPDRFSGSGSGTDFALTISRLEPEDFAVYYC QHYAGTPSNT
Beige background indicates a difference from human germline. Red font indicates amino acid differences from germline. Green font indicates primer-induced mutations.
Table S5. Competition of ADI-24247 and ADI-24314 with each other and with control mAbs.
ZV-67 4G2 ADI-24314 ADI-24247ADI-24247 1.0 1.4 1.5 53.2ADI-24314 1.0 2.3 61.8 1.4
IgG
Competitor Fab
Results are expressed as the fold reduction in antigen binding in the presence of saturating concentrations of competitor Fab relative to an antigen-only control.
Table S6. Binding properties of NS1-specific mAbs isolated from plasmablasts.
Clone Donor ZIKV NS1 DENV-1 NS1 DENV-2 NS1 DENV-3 NS1 DENV-4 NS1
ADI-32106 Donor 1 2.26E-08 N.B. N.B. N.B. N.B.
ADI-32111 Donor 1 2.51E-09 N.B. N.B. N.B. N.B.
ADI-32121 Donor 1 1.41E-08 N.B. N.B. N.B. N.B.
ADI-32123 Donor 1 7.72E-09 N.B. N.B. N.B. N.B.
ADI-32130 Donor 1 7.46E-09 N.B. N.B. N.B. N.B.
ADI-32140 Donor 1 3.95E-09 N.B. N.B. N.B. N.B.
ADI-32148 Donor 1 3.85E-09 N.B. N.B. N.B. N.B.
ADI-32155 Donor 1 1.72E-08 N.B. N.B. N.B. N.B.
ADI-32156 Donor 1 5.93E-09 N.B. N.B. N.B. N.B.
ADI-32160 Donor 1 1.62E-08 N.B. N.B. N.B. N.B.
ADI-32162 Donor 1 5.89E-09 N.B. N.B. N.B. N.B.
ADI-32163 Donor 1 1.71E-08 N.B. N.B. N.B. N.B.
ADI-32166 Donor 1 1.10E-09 N.B. N.B. N.B. N.B.
ADI-32167 Donor 1 6.67E-09 N.B. N.B. N.B. N.B.
ADI-32174 Donor 1 7.30E-09 N.B. N.B. N.B. N.B.
ADI-32178 Donor 1 1.85E-08 N.B. N.B. N.B. N.B.
ADI-32190 Donor 1 1.05E-09 N.B. N.B. N.B. N.B.
ADI-32192 Donor 1 8.73E-09 N.B. N.B. N.B. N.B.
ADI-32195 Donor 1 7.71E-09 N.B. N.B. N.B. N.B.
ADI-32196 Donor 1 1.32E-08 N.B. N.B. N.B. N.B.
ADI-32203 Donor 1 7.54E-09 N.B. N.B. N.B. N.B.
ADI-24189 Donor 2 1.05E-09 1.26E-09 N.B. 9.48E-10 6.81E-10
ADI-24199 Donor 2 1.27E-08 2.04E-09 N.B. 3.12E-09 N.B.
ADI-24201 Donor 2 7.16E-09 8.76E-10 N.B. 2.40E-09 N.B.
ADI-24204 Donor 2 2.64E-08 8.60E-10 N.B. 2.21E-08 N.B.
ADI-24205 Donor 2 2.74E-09 1.19E-09 N.B. 3.92E-09 N.B.
ADI-24206 Donor 2 1.87E-09 6.06E-10 N.B. 6.63E-10 N.B.
ADI-24207 Donor 2 3.21E-08 1.28E-09 N.B. 4.90E-09 N.B.
ADI-24215 Donor 2 5.48E-08 1.11E-08 N.B. 7.12E-09 1.33E-08
ADI-24265 Donor 3 7.01E-09 2.43E-09 N.B. 4.20E-09 N.B.
ADI-24287 Donor 3 1.18E-09 1.89E-09 N.B. 1.57E-09 N.B.
ADI-24290 Donor 3 1.12E-09 1.86E-09 N.B. 1.57E-09 N.B.
ADI-24291 Donor 3 1.11E-09 1.86E-09 N.B. 1.56E-09 N.B.
ADI-24305 Donor 4 1.35E-08 N.B. N.B. N.B. N.B.
ADI-24322 Donor 4 1.39E-09 N.B. N.B. N.B. N.B.
ADI-24324 Donor 4 4.97E-08 N.B. N.B. N.B. N.B.
ADI-24327 Donor 4 1.29E-07 N.B. N.B. N.B. N.B.
ADI-24337 Donor 4 8.31E-08 N.B. N.B. N.B. N.B.
ADI-24338 Donor 4 4.37E-09 N.B. N.B. N.B. N.B.
Apparent IgG KD (M)*
* N.B., non-binder; IgG KDs were calculated for MAbs with ForteBio binding responses >0.1 nm. MAbs with ForteBio binding responses <0.05 nm were designated as N.B.
Table S7. Neutralizing activity of selected plasmablast-derived mAbs.
Antibody Donor ZIKV-Para DENV-1 DENV-2 DENV-3 DENV-4 Epitope designationADI-24192 2 0.9 12 >3000 >3000 >3000 DIII (ZV-67
competitor)ADI-24227 2 2.1 9.0 >3000 >3000 >3000 DIII (ZV-67
competitor)ADI-24238 2 2.7 28 >3000 >3000 >3000 DIII (ZV-67
competitor)ADI-24220 2 452 275 2.5 21 20 FL (4G2-like)ADI-24211 2 595 819 7.0 >3000 20 FL (4G2-like)ADI-24241 2 608 >3000 >3000 >3000 >3000 UnknownADI-24203 2 >3000 >3000 >3000 >3000 >3000 UnknownADI-24236 2 >3000 >3000 >3000 >3000 >3000 UnknownADI-24226 2 >3000 1717 >3000 >3000 >3000 UnknownADI-24193 2 >3000 >3000 >3000 >3000 >3000 UnknownADI-24212 2 >3000 >3000 >3000 >3000 >3000 UnknownADI-24210 2 >3000 >3000 >3000 >3000 >3000 UnknownADI-24244 2 >3000 802 181 8.7 1954 ADI-24247ADI-24255 2 >3000 2.1 >3000 >3000 2299 DIII (ZV-67
competitor)ADI-24196 2 >3000 >3000 202 >3000 561 Whole-virus specificADI-24228 2 >3000 >3000 >3000 >3000 >3000 UnknownADI-24194 2 >3000 >3000 >3000 >3000 >3000 ADI-24314ADI-24249 2 >3000 >3000 203 >3000 1874 ADI-24247ADI-24195 2 >3000 5505 6.7 >3000 22 FL (4G2-like)ADI-24257 3 0.7 >3000 >3000 0.8 1437 ZV-67 competitorADI-24268 3 66 87 5.6 6.9 626 Whole-virus specificADI-24259 3 614 >3000 3.1 >3000 >3000 UnknownADI-24258 3 >3000 >3000 >3000 >3000 >3000 UnknownADI-24284 3 >3000 >3000 198 >3000 >3000 UnknownADI-24260 3 >3000 102 >3000 >3000 >3000 UnknownADI-24297 3 >3000 >3000 >3000 >3000 >3000 UnknownADI-24295 3 >3000 >3000 >3000 >3000 >3000 Whole-virus specificADI-24269 3 >3000 >3000 >3000 >3000 523 ADI-24314ADI-24294 3 >3000 4697 582 >3000 130 Whole-virus specificADI-24276 3 >3000 75 190 26 21 FL (4G2-like)ADI-24232 4 1.7 0.2 608 >3000 185 DIII (ZV-67
competitor)ADI-24367 4 5.9 8.2 >3000 196 >3000 ZV-67 competitorADI-24370 4 63 7.9 2.3 73 1494 UnknownADI-24361 4 72 87 8.0 >3000 62 UnknownADI-24373 4 182 24 8.6 32 >3000 UnknownADI-24326 4 192 612 310 >3000 64 UnknownADI-24343 4 215 >3000 >3000 >3000 >3000 Non WNV FLADI-24366 4 234 27 6.7 >3000 94 UnknownADI-24323 4 589 >3000 >3000 1515 >3000 Whole-virus specificADI-24318 4 627 28 75 24 >3000 Whole-virus specificADI-24375 4 >3000 >3000 >3000 >3000 >3000 UnknownADI-24307 4 >3000 >3000 >3000 >3000 >3000 UnknownADI-24359 4 >3000 >3000 >3000 >3000 >3000 UnknownADI-24310 4 >3000 >3000 >3000 >3000 >3000 UnknownADI-24374 4 >3000 23 25 >3000 1626 UnknownADI-24329 4 >3000 228 1125 >3000 >3000 UnknownADI-24320 4 >3000 >3000 159 522 4683 Unknown
Neutralization IC50 (ng/ml)
Table S8. Binding properties of ZIKV E-specific mAbs isolated from memory B cells.
Clone Donor DIII ZIKV E DENV-1 E DENV-2 E DENV-3 E DENV-4 E ZV-67 4G2 ADI-24314 ADI-24247 ADI-30023 ADI-30056 Epitope designation% Neutralization of
ZIKV-Para at 1 ug/ml
ADI-30008 3 N.B. 9.51E-10 5.98E-10 6.66E-10 4.38E-10 4.33E-10 0.9 67.8 N/A N/A N/A N/A 4G2 0
ADI-30013 3 N.B. 1.27E-09 6.59E-10 7.72E-10 4.87E-10 4.82E-10 1.0 47.9 N/A N/A N/A N/A 4G2 0
ADI-30053 3 N.B. 1.30E-09 1.09E-09 9.81E-10 5.43E-10 5.96E-10 1.2 44.9 N/A N/A N/A N/A 4G2 0
ADI-30049 3 N.B. 1.52E-09 9.35E-10 1.12E-09 6.88E-10 5.88E-10 0.9 39.3 N/A N/A N/A N/A 4G2 0
ADI-30057 3 N.B. 1.64E-09 1.17E-09 1.56E-09 7.99E-10 9.17E-10 0.9 38.4 N/A N/A N/A N/A 4G2 0
ADI-30024 3 N.B. 7.60E-10 6.78E-10 5.47E-10 3.30E-10 2.91E-10 0.9 30.4 N/A N/A N/A N/A 4G2 0
ADI-30001 3 N.B. 1.24E-09 7.22E-10 7.61E-10 4.35E-10 4.54E-10 1.1 27.1 N/A N/A N/A N/A 4G2 50
ADI-29999 3 N.B. 1.44E-09 7.41E-10 7.96E-10 4.66E-10 4.63E-10 1.1 25.4 N/A N/A N/A N/A 4G2 0
ADI-30059 3 N.B. 1.19E-09 8.22E-10 9.35E-10 5.02E-10 4.75E-10 0.9 24.4 N/A N/A N/A N/A 4G2 0
ADI-29998 3 N.B. 1.09E-09 7.73E-10 8.93E-10 5.39E-10 6.05E-10 1.2 24.0 N/A N/A N/A N/A 4G2 0
ADI-30014 3 N.B. 1.53E-09 7.76E-10 9.58E-10 8.48E-10 5.04E-10 0.9 23.8 N/A N/A N/A N/A 4G2 49
ADI-30017 3 N.B. 1.10E-09 9.59E-10 7.30E-10 4.41E-10 4.21E-10 0.9 23.2 N/A N/A N/A N/A 4G2 0
ADI-30048 3 N.B. 1.52E-09 3.50E-09 3.23E-09 1.75E-09 1.80E-09 0.8 21.2 N/A N/A N/A N/A 4G2 0
ADI-30043 3 N.B. 1.28E-09 1.02E-09 9.21E-10 4.99E-10 5.23E-10 0.8 21.0 N/A N/A N/A N/A 4G2 0
ADI-30020 3 N.B. 1.39E-09 8.41E-10 8.90E-10 5.32E-10 4.85E-10 0.9 19.6 N/A N/A N/A N/A 4G2 0
ADI-30019 3 N.B. 9.23E-10 1.15E-09 1.03E-09 4.89E-10 4.26E-10 0.8 18.1 N/A N/A N/A N/A 4G2 0
ADI-30009 3 N.B. 1.12E-09 7.34E-10 8.19E-10 5.62E-10 6.01E-10 1.0 18.1 N/A N/A N/A N/A 4G2 0
ADI-29995 3 N.B. 1.08E-09 6.82E-10 8.22E-10 4.99E-10 5.45E-10 1.0 16.9 N/A N/A N/A N/A 4G2 0
ADI-29992 3 N.B. 1.14E-09 8.98E-10 7.80E-10 4.54E-10 4.45E-10 0.8 16.2 N/A N/A N/A N/A 4G2 0
ADI-29996 3 N.B. 1.14E-09 1.12E-09 5.77E-10 3.95E-10 3.92E-10 1.0 15.4 N/A N/A N/A N/A 4G2 0
ADI-30010 3 N.B. 1.18E-09 6.54E-10 6.40E-10 4.48E-10 4.15E-10 1.2 14.7 N/A N/A N/A N/A 4G2 0
ADI-30063 3 N.B. 8.90E-08 N.B. N.B. N.B. N.B. 1.0 14.2 N/A N/A N/A N/A 4G2 44
ADI-30044 3 N.B. 3.33E-08 N.B. N.B. N.B. N.B. 1.1 9.7 N/A N/A N/A N/A 4G2 0
ADI-30069 3 N.B. 3.33E-08 1.19E-07 5.58E-08 3.87E-08 6.32E-08 0.7 8.0 N/A N/A N/A N/A 4G2 0
ADI-30042 3 N.B. 9.05E-09 N.B. N.B. N.B. N.B. 1.1 6.9 N/A N/A N/A N/A 4G2 0
ADI-30050 3 N.B. 2.00E-08 N.B. N.B. N.B. N.B. 1.0 6.5 N/A N/A N/A N/A 4G2 62
ADI-30002 3 N.B. 8.26E-09 N.B. N.B. N.B. N.B. 0.8 5.4 N/A N/A N/A N/A 4G2 0
ADI-30046 3 N.B. 7.27E-09 N.B. N.B. N.B. N.B. 0.8 4.3 N/A N/A N/A N/A 4G2 99
ADI-30026 3 N.B. 1.16E-08 7.79E-10 6.09E-10 4.31E-10 3.67E-10 1.4 3.9 N/A N/A N/A N/A 4G2 0
ADI-30077 3 N.B. 1.90E-09 N.B. N.B. N.B. N.B. 1.2 3.8 N/A N/A N/A N/A 4G2 0
ADI-30052 3 N.B. 5.60E-08 N.B. N.B. N.B. N.B. 1.1 3.8 N/A N/A N/A N/A 4G2 97
ADI-29993 3 N.B. 2.87E-09 3.52E-09 1.06E-09 1.03E-09 7.59E-10 1.0 1.4 1.4 21.0 N/A N/A ADI-24247 0
ADI-30015 3 N.B. 2.73E-09 1.51E-09 9.14E-10 7.51E-10 6.06E-10 1.1 1.7 1.3 6.6 N/A N/A ADI-24247 0
ADI-30066 3 N.B. 1.81E-09 1.94E-09 1.01E-09 8.09E-10 2.69E-09 1.0 2.0 35.0 1.6 N/A N/A ADI-24314 0
ADI-30021 3 N.B. 1.50E-09 N.B. N.B. N.B. N.B. 1.0 2.8 5.8 1.5 N/A N/A ADI-24314 86
ADI-30030 3 N.B. 1.56E-08 N.B. N.B. N.B. N.B. 0.8 1.5 1.5 3.5 9.3 2.5 ADI-30023 0
ADI-30023 3 N.B. 1.89E-09 N.B. N.B. N.B. N.B. 1.1 1.9 2.3 1.7 9.9 2.0 ADI-30023 87
ADI-30032 3 N.B. 2.33E-09 N.B. N.B. N.B. N.B. 1.2 2.0 1.3 1.7 17.3 2.0 ADI-30023 100
ADI-30089 3 N.B. 3.10E-08 N.B. N.B. N.B. N.B. 1.1 2.4 2.1 1.5 2.0 16.9 ADI-30056 53
ADI-30038 3 N.B. 2.21E-09 N.B. N.B. N.B. N.B. 0.9 2.3 1.5 1.5 1.7 24.9 ADI-30056 77
ADI-30083 3 N.B. 3.49E-09 N.B. N.B. N.B. N.B. 1.1 2.0 1.6 1.5 1.7 28.6 ADI-30056 100
ADI-30022 3 N.B. 3.62E-09 N.B. N.B. N.B. N.B. 1.5 1.7 3.4 1.5 4.2 7.9 ADI-30056 0
ADI-30033 3 N.B. 3.56E-09 N.B. N.B. N.B. N.B. 1.0 1.3 1.6 1.4 1.9 29.8 ADI-30056 0
ADI-30062 3 N.B. 1.87E-09 N.B. N.B. N.B. N.B. 1.0 3.0 1.5 1.4 1.7 39.4 ADI-30056 46
ADI-29994 3 N.B. 1.77E-09 N.B. N.B. N.B. N.B. 1.0 1.9 1.5 1.4 1.7 12.2 ADI-30056 100
ADI-30075 3 N.B. 3.00E-09 N.B. N.B. N.B. N.B. 1.0 1.6 1.2 1.4 1.5 44.2 ADI-30056 100
ADI-30056 3 N.B. 1.84E-09 N.B. N.B. N.B. N.B. 1.0 1.9 1.6 1.4 1.5 57.8 ADI-30056 100
ADI-30035 3 N.B. 1.13E-09 N.B. N.B. N.B. 3.54E-08 1.1 2.2 1.3 1.4 1.4 14.1 ADI-30056 0
ADI-30006 3 N.B. 2.42E-09 N.B. N.B. N.B. N.B. 0.9 1.4 1.7 1.4 1.7 15.6 ADI-30056 100
ADI-30061 3 N.B. 5.78E-09 N.B. N.B. N.B. N.B. 1.0 1.7 2.0 1.4 3.4 16.0 ADI-30056 100
ADI-30003 3 N.B. 2.02E-09 N.B. N.B. N.B. 9.69E-08 0.9 1.4 3.6 1.3 1.4 26.1 ADI-30056 30
ADI-30037 3 N.B. 9.57E-10 N.B. N.B. N.B. N.B. 0.9 1.9 1.5 1.3 1.4 10.4 ADI-30056 0
ADI-30031 3 N.B. 4.86E-09 N.B. N.B. N.B. N.B. 0.9 1.7 1.5 1.3 1.6 51.9 ADI-30056 100
ADI-30025 3 N.B. 1.75E-09 N.B. N.B. N.B. N.B. 0.8 1.6 1.1 1.2 1.5 50.2 ADI-30056 95
ADI-30072 3 N.B. 6.19E-08 N.B. N.B. N.B. N.B. 1.0 2.0 2.4 1.2 1.3 4.3 ADI-30056 0
ADI-30039 3 N.B. 1.02E-09 N.B. N.B. N.B. N.B. 0.8 1.4 1.3 1.2 1.5 69.8 ADI-30056 97
ADI-30055 3 N.B. 7.20E-10 N.B. N.B. N.B. N.B. 0.9 2.0 1.6 1.2 1.3 36.7 ADI-30056 91
ADI-30045 3 N.B. 2.82E-09 N.B. N.B. N.B. N.B. 0.9 2.5 1.2 1.2 1.4 61.8 ADI-30056 100
ADI-29997 3 N.B. 8.12E-09 N.B. N.B. N.B. N.B. 0.8 2.1 1.7 1.2 1.4 8.1 ADI-30056 100
ADI-30000 3 N.B. 2.38E-09 N.B. N.B. N.B. N.B. 0.9 1.8 1.4 1.2 1.6 27.4 ADI-30056 98
ADI-30007 3 N.B. 1.35E-08 N.B. N.B. N.B. N.B. 1.1 1.5 3.0 1.1 1.3 3.7 ADI-30056 98
ADI-30034 3 N.B. 2.96E-09 N.B. N.B. N.B. N.B. 1.0 2.1 1.6 1.1 1.4 3.2 ADI-30056 97
ADI-30029 3 N.B. 1.73E-08 N.B. N.B. N.B. N.B. 0.8 2.3 0.9 1.1 1.4 4.6 ADI-30056 100
ADI-30005 3 N.B. 4.38E-09 N.B. N.B. N.B. N.B. 0.7 1.7 0.9 1.0 1.1 6.1 ADI-30056 72
ADI-30016 3 N.B. 2.09E-08 N.B. N.B. N.B. N.B. 0.8 1.7 1.1 1.0 3.0 10.1 ADI-30056 60
ADI-30071 3 N.B. 1.22E-08 N.B. N.B. N.B. N.B. 0.8 2.4 0.8 0.8 1.2 9.5 ADI-30056 100
ADI-30040 3 N.B. 4.35E-08 N.B. N.B. N.B. N.B. 1.0 1.2 1.0 0.8 1.5 4.1 ADI-30056 99
ADI-30087 3 N.B. 1.04E-08 N.B. N.B. N.B. N.B. 1.0 1.4 0.5 0.6 1.3 7.7 ADI-30056 98
ADI-30058 3 N.B. 1.35962E-09 1.838E-09 2.243E-09 1.026E-09 7.245E-10 N/A N/A N/A N/A N/A N/A NC 0
ADI-30064 3 N.B. 4.22E-09 N.B. N.B. N.B. N.B. N/A N/A N/A N/A N/A N/A NC 51
ADI-30065 3 N.B. 8.59E-08 N.B. N.B. N.B. N.B. N/A N/A N/A N/A N/A N/A NC 96
ADI-30070 3 N.B. 7.15E-08 N.B. N.B. N.B. N.B. N/A N/A N/A N/A N/A N/A NC 0
ADI-30081 3 N.B. 7.76E-08 N.B. N.B. N.B. N.B. N/A N/A N/A N/A N/A N/A NC 97
ADI-30086 3 N.B. 3.84E-08 N.B. N.B. N.B. N.B. N/A N/A N/A N/A N/A N/A NC 0
ADI-30085 3 4.574E-09 5.04E-09 N.B. N.B. N.B. N.B. 9.6 2.3 N/A N/A N/A N/A ZV67 0
ADI-30079 3 6.299E-09 9.24E-09 N.B. N.B. N.B. N.B. 10.0 2.0 N/A N/A N/A N/A ZV67 0
ADI-30028 3 8.957E-10 2.18E-09 N.B. N.B. N.B. N.B. 14.6 1.5 N/A N/A N/A N/A ZV67 95
ADI-30054 3 5.714E-10 1.57E-09 N.B. N.B. N.B. N.B. 13.2 1.4 N/A N/A N/A N/A ZV67 96
ADI-30051 3 N.B. 6.24E-09 N.B. N.B. N.B. N.B. 2.2 1.0 N/A N/A N/A N/A ZV67 97
ADI-30012 3 6.727E-10 2.14E-09 N.B. N.B. N.B. N.B. 2.5 2.0 1.6 1.6 1.5 1.7 ZV67 66
ADI-30152 4 N.B. 7.82E-10 5.84E-10 6.51E-10 3.62E-10 3.66E-10 1.0 85.9 N/A N/A N/A N/A 4G2 0
ADI-30168 4 N.B. 1.08E-09 1.06E-09 1.91E-09 1.15E-09 9.16E-10 0.9 81.9 N/A N/A N/A N/A 4G2 0
Apparent IgG KD (M)* Antibody competition (Fold reduction in antigen binding in the presence of competitor Fab)#
Clone Donor DIII ZIKV E DENV-1 E DENV-2 E DENV-3 E DENV-4 E ZV-67 4G2 ADI-24314 ADI-24247 ADI-30023 ADI-30056 Epitope designation% Neutralization of
ZIKV-Para at 1 ug/ml
Apparent IgG KD (M)* Antibody competition (Fold reduction in antigen binding in the presence of competitor Fab)#
ADI-30141 4 N.B. 9.11E-10 9.83E-10 1.18E-09 9.68E-10 6.61E-10 0.9 78.7 N/A N/A N/A N/A 4G2 0ADI-30149 4 N.B. 1.23E-09 1.15E-09 8.17E-10 6.82E-10 4.92E-10 1.0 75.8 N/A N/A N/A N/A 4G2 0ADI-30150 4 N.B. 6.69E-10 3.81E-10 5.20E-10 2.34E-10 2.39E-10 0.9 73.9 N/A N/A N/A N/A 4G2 0ADI-30151 4 N.B. 1.27E-09 9.27E-10 9.69E-10 5.18E-10 5.62E-10 1.2 66.2 N/A N/A N/A N/A 4G2 0ADI-30154 4 N.B. 1.01E-09 7.15E-10 7.36E-10 4.01E-10 3.85E-10 1.1 57.7 N/A N/A N/A N/A 4G2 0ADI-30148 4 N.B. 1.08E-09 9.29E-10 8.30E-10 6.06E-10 5.17E-10 1.2 57.5 N/A N/A N/A N/A 4G2 0ADI-30180 4 N.B. 9.87E-10 6.52E-10 6.23E-10 5.17E-10 5.11E-10 1.3 56.9 N/A N/A N/A N/A 4G2 0ADI-30181 4 N.B. 1.21E-09 2.64E-09 2.46E-09 1.77E-09 1.49E-09 1.0 53.4 N/A N/A N/A N/A 4G2 0ADI-30155 4 N.B. 1.25E-09 1.59E-09 1.15E-09 5.49E-10 4.99E-10 1.1 53.0 N/A N/A N/A N/A 4G2 0ADI-30177 4 N.B. 1.04E-09 7.86E-10 6.84E-10 3.83E-10 3.39E-10 1.3 44.0 N/A N/A N/A N/A 4G2 0ADI-30142 4 N.B. 1.03E-09 7.36E-10 7.78E-10 4.14E-10 3.71E-10 1.2 42.5 N/A N/A N/A N/A 4G2 0ADI-30153 4 N.B. 1.34E-09 8.70E-10 9.19E-10 5.21E-10 5.68E-10 1.3 41.3 N/A N/A N/A N/A 4G2 0ADI-30107 4 N.B. 1.49E-09 8.65E-10 7.86E-10 5.82E-10 5.18E-10 1.2 39.9 N/A N/A N/A N/A 4G2 0ADI-30167 4 N.B. 1.55E-09 2.28E-09 2.42E-09 1.83E-09 1.47E-09 1.1 37.3 N/A N/A N/A N/A 4G2 0ADI-30162 4 N.B. 1.48E-09 1.30E-09 1.26E-09 1.14E-09 8.36E-10 1.2 35.2 N/A N/A N/A N/A 4G2 0ADI-30116 4 N.B. 1.16E-09 9.33E-10 9.31E-10 6.95E-10 3.92E-10 1.2 32.5 N/A N/A N/A N/A 4G2 0ADI-30169 4 N.B. 1.13E-09 1.09E-09 7.52E-10 4.73E-10 4.66E-10 1.3 30.0 N/A N/A N/A N/A 4G2 0ADI-30132 4 N.B. 1.34E-09 9.10E-10 9.51E-10 9.03E-10 5.15E-10 1.2 29.5 N/A N/A N/A N/A 4G2 0ADI-30166 4 N.B. 1.14E-09 1.10E-09 8.46E-10 5.89E-10 9.50E-10 1.3 27.5 N/A N/A N/A N/A 4G2 0ADI-30114 4 N.B. 1.27E-09 6.21E-10 6.81E-10 5.42E-10 3.42E-10 1.3 26.9 N/A N/A N/A N/A 4G2 0ADI-30175 4 N.B. 1.12E-09 8.26E-10 7.56E-10 4.82E-10 4.26E-10 1.0 26.4 N/A N/A N/A N/A 4G2 0ADI-30119 4 N.B. 1.94E-09 2.80E-09 3.59E-09 1.90E-09 2.08E-09 1.3 26.0 N/A N/A N/A N/A 4G2 0ADI-30156 4 N.B. 4.80E-10 6.00E-10 3.92E-10 3.95E-10 2.00E-10 0.9 24.8 N/A N/A N/A N/A 4G2 0ADI-30130 4 N.B. 1.10E-09 1.12E-09 8.20E-10 6.83E-10 3.79E-10 1.1 23.3 N/A N/A N/A N/A 4G2 0ADI-30125 4 N.B. 1.00E-09 4.83E-10 7.17E-10 4.89E-10 3.39E-10 1.3 23.2 N/A N/A N/A N/A 4G2 0ADI-30124 4 N.B. 1.12E-09 5.77E-10 6.81E-10 4.62E-10 3.56E-10 1.2 21.5 N/A N/A N/A N/A 4G2 0ADI-30134 4 N.B. 1.29E-09 7.07E-10 7.84E-10 4.46E-10 4.24E-10 1.7 21.5 N/A N/A N/A N/A 4G2 0ADI-30147 4 N.B. 1.45E-09 9.58E-10 9.59E-10 5.44E-10 5.37E-10 1.1 19.5 N/A N/A N/A N/A 4G2 0ADI-30118 4 N.B. 1.40E-09 1.86E-09 2.44E-09 1.36E-09 1.41E-09 1.2 17.5 N/A N/A N/A N/A 4G2 0ADI-30133 4 N.B. 9.41E-09 2.36E-08 5.32E-08 2.48E-08 2.30E-08 0.9 14.9 N/A N/A N/A N/A 4G2 0ADI-30159 4 N.B. 1.49E-09 9.66E-10 9.83E-10 5.92E-10 6.31E-10 1.0 14.5 N/A N/A N/A N/A 4G2 0ADI-30172 4 N.B. 1.77E-08 1.22E-09 9.08E-10 5.61E-10 4.71E-10 0.9 4.4 N/A N/A N/A N/A 4G2 0ADI-30121 4 N.B. 9.04E-10 N.B. N.B. 4.41E-08 3.86E-10 1.2 4.3 N/A N/A N/A N/A 4G2 0ADI-30111 4 N.B. 1.22E-09 6.57E-10 7.87E-10 4.56E-10 4.10E-10 1.1 3.2 N/A N/A N/A N/A 4G2 0ADI-30170 4 N.B. 2.22E-09 N.B. 9.55E-08 N.B. 7.39E-10 1.1 1.5 1.0 29.6 N/A N/A ADI-24247 0ADI-30109 4 N.B. 2.14E-09 N.B. N.B. N.B. 2.55E-09 1.4 1.2 0.9 22.1 N/A N/A ADI-24247 0ADI-30127 4 N.B. 2.71E-09 8.50E-09 1.49E-09 1.21E-08 9.42E-10 1.3 1.6 1.2 6.8 N/A N/A ADI-24247 0ADI-30136 4 N.B. 5.02E-09 8.78E-10 7.65E-10 5.09E-10 4.06E-10 1.0 2.4 42.7 3.9 N/A N/A ADI-24314 0ADI-30144 4 N.B. 2.32E-09 1.17E-08 2.88E-08 4.77E-09 1.13E-08 1.1 2.6 38.4 2.6 N/A N/A ADI-24314 0ADI-30122 4 N.B. 1.88E-09 8.47E-10 7.55E-10 4.72E-10 4.52E-10 1.1 1.7 24.6 2.1 N/A N/A ADI-24314 0ADI-30182 4 N.B. 1.18E-09 6.85E-10 5.56E-10 3.95E-10 3.43E-10 1.1 1.5 13.7 1.6 N/A N/A ADI-24314 0ADI-30123 4 N.B. 1.03E-09 9.84E-10 5.59E-10 3.57E-10 3.52E-10 1.3 3.0 17.1 1.5 N/A N/A ADI-24314 0ADI-30163 4 N.B. 1.59E-09 8.93E-10 7.52E-10 4.92E-10 4.30E-10 0.9 1.7 47.5 1.4 N/A N/A ADI-24314 0ADI-30098 4 N.B. 1.03E-09 N.B. 2.42E-10 N.B. 2.01E-10 0.9 2.8 4.3 1.2 N/A N/A ADI-24314 0ADI-30131 4 N.B. 4.49E-09 N.B. N.B. N.B. N.B. 1.2 2.1 1.6 1.8 4.1 2.0 ADI-30023 73ADI-30164 4 N.B. 1.30E-09 N.B. N.B. N.B. N.B. 1.1 1.5 1.1 1.4 10.2 1.7 ADI-30023 64ADI-30093 4 N.B. 3.09E-09 N.B. N.B. N.B. N.B. 1.4 1.9 2.0 1.8 2.3 9.0 ADI-30056 83ADI-30174 4 N.B. 1.51E-09 N.B. N.B. N.B. N.B. 1.2 2.5 2.3 1.8 1.8 4.4 ADI-30056 0ADI-30120 4 N.B. 9.79E-10 9.52E-08 N.B. 1.17E-09 4.23E-10 1.3 2.3 1.4 1.7 1.6 23.2 ADI-30056 0ADI-30158 4 N.B. 1.53E-09 N.B. N.B. N.B. 6.39E-09 0.9 1.7 1.4 1.6 1.7 30.1 ADI-30056 0ADI-30110 4 N.B. 1.21E-09 N.B. N.B. 8.49E-08 4.30E-10 1.3 2.8 1.2 1.5 1.5 14.4 ADI-30056 0ADI-30137 4 N.B. 6.14E-09 N.B. N.B. N.B. N.B. 1.0 1.9 1.2 1.5 1.5 14.7 ADI-30056 74ADI-30135 4 N.B. 1.44E-09 N.B. N.B. N.B. N.B. 1.2 1.8 1.4 1.5 1.5 8.0 ADI-30056 77ADI-30084 4 N.B. 4.88E-09 N.B. N.B. N.B. N.B. 1.0 3.7 1.1 1.3 1.5 32.6 ADI-30056 95ADI-30097 4 N.B. 1.50E-08 N.B. N.B. N.B. N.B. 1.1 0.9 1.1 1.3 1.6 30.0 ADI-30056 100ADI-30091 4 N.B. 5.22E-09 N.B. N.B. N.B. N.B. 1.0 2.0 1.2 1.3 1.4 43.1 ADI-30056 91ADI-30101 4 N.B. 7.99E-09 N.B. N.B. N.B. N.B. 1.0 2.8 1.1 1.3 1.5 27.0 ADI-30056 97ADI-30100 4 N.B. 5.03E-09 N.B. N.B. N.B. N.B. 1.1 2.4 1.2 1.2 1.4 11.3 ADI-30056 99ADI-30171 4 N.B. 1.15E-09 N.B. N.B. 5.20E-08 2.59E-09 1.0 2.6 0.8 1.0 1.5 25.2 ADI-30056 0ADI-30103 4 N.B. 1.16E-08 N.B. N.B. N.B. N.B. N/A N/A N/A N/A N/A N/A NC 89ADI-30139 4 N.B. 4.72E-08 3.08E-07 N.B. 6.31E-08 1.49E-07 N/A N/A N/A N/A N/A N/A NC 0ADI-30179 4 N.B. 2.15E-08 N.B. N.B. N.B. 1.46E-07 N/A N/A N/A N/A N/A N/A NC 0ADI-30117 4 N.B. 3.35E-09 7.42E-09 N.B. N.B. N.B. 1.3 3.4 2.3 1.8 3.0 2.9 unknown 72ADI-30145 4 N.B. 1.87E-09 N.B. N.B. N.B. N.B. 1.0 1.7 1.5 1.8 1.6 1.6 unknown 63ADI-30126 4 6.810E-10 1.85E-09 N.B. N.B. N.B. N.B. 2.9 1.4 1.1 1.7 1.5 1.5 DIII 0ADI-30157 4 3.231E-09 1.70E-09 N.B. N.B. N.B. N.B. 2.7 1.5 0.9 1.6 1.5 1.4 DIII 96ADI-30160 4 N.B. 7.66E-09 N.B. N.B. N.B. N.B. 1.1 2.0 1.2 1.3 1.7 2.6 unknown 65ADI-30128 4 N.B. 1.42E-08 N.B. N.B. N.B. N.B. 1.1 2.9 1.3 1.1 1.6 2.1 unknown 0ADI-30074 4 2.200E-09 3.52E-09 N.B. N.B. N.B. N.B. 13.7 2.1 N/A N/A N/A N/A ZV-67 0ADI-30060 4 2.104E-09 3.60E-09 N.B. N.B. N.B. N.B. 7.2 1.9 N/A N/A N/A N/A ZV-67 0ADI-30112 4 N.B. 6.72E-08 N.B. N.B. N.B. N.B. 6.7 1.9 N/A N/A N/A N/A ZV-67 77ADI-30095 4 1.384E-09 7.26E-09 N.B. N.B. N.B. N.B. 7.3 1.7 N/A N/A N/A N/A ZV-67 0ADI-30140 4 6.081E-10 2.26E-09 N.B. N.B. N.B. N.B. 11.0 1.4 N/A N/A N/A N/A ZV-67 100
* N.B., non-binder; W.B., weak binder. IgG KDs were calculated for MAbs with ForteBio binding responses >0.1 nm. MAbs with ForteBio binding responses <0.05 nm were designated as N.B.; IgGs with ForteBio binding responses between 0.05 and 0.1 nm were designated as W.B. # Results are expressed as the fold reduction in antigen binding in the presence of saturating concentrations of competitor Fab relative to an antigen-only control.
Table S9. Clonal lineages shared between plasmablast and memory B cell-derived antibodies.
Donor B cell subset Lineage # VH germline HC FR1 HC CDR1 HC FR2 HC CDR2 HC FR3 HC CDR3 VL germline* LC FR1 LC CDR1 LC FR2 LC CDR2 LC FR3 LC CDR3
Donor 3 memory 1 VH3-30-3 EVQLVESGGGVVQPGRSLRLSCVASG FTFSDNALH WVRQAPGKALEWVA VTSYDGSSKFYADSVKG RFTISRDTSKNTLYLQMNSLRPEDTATYYC ASRGRLPALAEYFQH VL1-40 QSVLTQPPSVSGAPGQKVTISC TGSSSNIGTHFDVH WYQQLPGTAPKLLIY GNTNRPS GVPDRFSGSKSGTSASLAITGLQAEDEADYYC QSYDSSLTAVVDonor 3 memory 1 VH3-30-3 QVQLQESGGGVVQPGRSLRLSCAASG FTFSDNAMH WVRQAPGKALEWVA VTSYDGSETYYADSVKG RFTISRDSSKNMLYLQMNSLSTEDTAVYYC ASRGVLPALAEYFQH VL1-40 QSVLTQPPSVSGAPGRRVTISC AGSSSNIGAGYDVH WYQQLPGTAPKLLIY GNSNRPS GVPDRFSGSKSGTSASLAITGLQAEDEADYYC QSYDSSLTGVVDonor 3 plasmablast 1 VH3-30-3 EVQLVESGGGVVQPGRSLRLSCAASG FTFSDNAMH WVRQAPGKALEWVA VASYDGTQKYYADSVKG RFTISRDTSKNTLYLQMNSLKPEDTAIYYC ASRGLLPALAEYFQY VL1-40 QSVLTQPPSVSGAPGQRVTISC TGSNSNIGAHFDVH WYQQLPGTAPKLLVY GNSNRPS GVPDRFSGSKSGTSASLAIAGLQAEDEADYYC QSYDSSLTAVVDonor 3 memory 2 VH3-7 EVQLLESGGDLVQPGGSLRISCAASK FTFSNYLMS WVRQAPGKGLEWVA NIRQDGTETYYVDSVKG RFTISRHNAKNSLYLQMSSLRVEDTAVYYC ARNRRDCSSGSCYDSGVDFDM VL2-23 QPVLTQPASVSGSPGQSITISC TGTNSDVDNYNLVS WYQQHPGKAPKLMIY EVTKRPS GISNRFSGSKSGSTASLTISGLQAEDEADYYC CSYGRSSALDonor 3 plasmablast 2 VH3-7 EVQLVESGGDLVQPGGSLRLSCAASK FTFSNYLMS WVRQAPGKGLEWVA NIRQDGTEKYYVDSVKG RFTISRDNAKSSVYLQMNSLRLEDTAVYYC ARNRRDCSSDHCYDSGVDFDL N/A N/A N/A N/A N/A N/A N/ADonor 3 plasmablast 2 VH3-7 EVQLVESGGGLVQPGGSLRLSCAGSG FTFSSFLMS WVRQPPGKGLEWVA NIRQDGIEKYYLDSVRG RFTISRDNAKNSLSLQMNSLRADDTAVYFC ARNRRDCGGRSCYDSGIDFDV N/A N/A N/A N/A N/A N/A N/ADonor 3 memory 3 VH3-30 EVQLLESGGRVVQPGRSLRLSCAASG FSFGTDAMH WVRQAPGKGLEWVA VISYDGTSTFYADSVKG RFTISRDNSKNTLYLQMNSLRVEDTAIYFC AREGHCTGATCYSTSFDF VL1-40 QSALIQPPSVSGAPGQRVTISC TGSSPNIGPGYDVH WYQQLPGKAPKLLIY GNTNRPS GVPDRFSGSKSGTSASLAISALQAEDEADYYC QSYDSSLSAVIDonor 3 plasmablast 3 VH3-30 EVQLVESGGAVVQPGRSLRLSCAASG FTFSNDAMH WVRQAPGKGLEWVA VISLDGRQKYYSDSVKG RFTISRDNSKNTVFLQMNSLRVEDTATYYC AREGHCTGATCYSTSFDF VL1-40 QSVLTQPPSVSGAPGQRVTISC TGSSPNIGPGYDVH WYQQIPGTAPKLLIY GNTNRPS GVPDRFSGSKSGTSASLAITGLQPEDEADYYC QTYDRTLNGVV
Beige background indicates a difference from human germline. Red font indicates amino acid differences from germline. *N/A, not applicable due to lack of light chain amplification
Table S10. Neutralizing activity of selected memory B cell-derived mAbs.
Clone ZIKV-Para DENV-1 DENV-2 DENV-3 DENV-4ADI-30001 1082 132 110 1429 8485ADI-30117 300 145 8367 13563 12727ADI-30114 3142 363 110 1148 1379ADI-29999 3381 363 260 289 2624ADI-29993 4235 377 4588 7586 3532ADI-30124 1264 382 113 1327 1245ADI-30017 3176 391 100 442 3633ADI-29992 3245 396 153 1327 3027ADI-30132 1150 415 159 3525 4238ADI-30116 134 433 51 1046 1379ADI-30130 277 433 113 315 1278ADI-30125 414 447 143 1250 12424ADI-30127 2254 471 1288 391 15152ADI-29998 9436 1019 156 4210 3532ADI-30107 3210 1061 156 842 4137ADI-30015 1275 1075 2879 8276 3935ADI-30111 417 1201 58 1046 8182ADI-30009 4064 1201 110 3139 7879ADI-30134 1275 1229 143 3908 1446ADI-30043 8410 1410 320 3062 11212ADI-30059 8923 1424 260 1403 4137ADI-30122 9077 4314 489 944 3636ADI-30110 1002 13333 4674 1148 3229ADI-30109 140 16352 10526 11264 11515ADI-30056 1.8 N/A N/A N/A N/AADI-30031 5.2 N/A N/A N/A N/AADI-30000 7.7 N/A N/A N/A N/AADI-30157 11.6 N/A N/A N/A N/AADI-29997 15 N/A N/A N/A N/AADI-30006 30 N/A N/A N/A N/AADI-30007 34 N/A N/A N/A N/AADI-30140 35.0 N/A N/A N/A N/AADI-29994 45 N/A N/A N/A N/AADI-30097 45.3 N/A N/A N/A N/AADI-30032 64 N/A N/A N/A N/AADI-30025 87 N/A N/A N/A N/AADI-30054 90 N/A N/A N/A N/AADI-30028 94 N/A N/A N/A N/AADI-30131 98.2 N/A N/A N/A N/AADI-30100 103 N/A N/A N/A N/AADI-30084 105 N/A N/A N/A N/AADI-30034 125 N/A N/A N/A N/AADI-30021 133 N/A N/A N/A N/AADI-30016 138 N/A N/A N/A N/AADI-30101 142 N/A N/A N/A N/A
Neutralization IC50 (ng/ml)
Clone ZIKV-Para DENV-1 DENV-2 DENV-3 DENV-4Neutralization IC50 (ng/ml)
ADI-30135 159 N/A N/A N/A N/AADI-30093 204 N/A N/A N/A N/AADI-30091 223 N/A N/A N/A N/AADI-30112 309 N/A N/A N/A N/AADI-30023 321 N/A N/A N/A N/AADI-30137 327 N/A N/A N/A N/AADI-30005 351 N/A N/A N/A N/AADI-30160 399 N/A N/A N/A N/AADI-30038 405 N/A N/A N/A N/AADI-30012 497 N/A N/A N/A N/AADI-30062 660 N/A N/A N/A N/AADI-30022 1133 N/A N/A N/A N/AADI-30126 1178 N/A N/A N/A N/AADI-30174 2436 N/A N/A N/A N/AADI-30037 2463 N/A N/A N/A N/AADI-30002 2999 N/A N/A N/A N/AADI-30128 3079 N/A N/A N/A N/AADI-30030 3909 N/A N/A N/A N/AADI-30060 5301 N/A N/A N/A N/AADI-30033 8916 N/A N/A N/A N/AADI-30074 9960 N/A N/A N/A N/AADI-30079 10361 N/A N/A N/A N/AADI-30095 12129 N/A N/A N/A N/AADI-30085 13414 N/A N/A N/A N/A