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1.
Cell Reports ; : 110812, 2022.
Article in English | ScienceDirect | ID: covidwho-1803708

ABSTRACT

SARS-CoV-2 neutralizing monoclonal antibodies (mAbs) can reduce the risk of hospitalization from COVID-19 when administered early. However, SARS-CoV-2 variants of concern (VOC) have negatively impacted the therapeutic use of some authorized mAbs. Using a high throughput B-cell screening pipeline, we isolate LY-CoV1404 (bebtelovimab), a highly potent SARS-CoV-2 spike glycoprotein receptor binding domain (RBD)-specific antibody. LY-CoV1404 potently neutralizes authentic SARS-CoV-2, B.1.1.7, B.1.351 and B.1.617.2. In pseudovirus neutralization studies, LY-CoV1404 potently neutralizes variants including B.1.1.7, B.1.351, B.1.617.2, B.1.427/B.1.429, P.1, B.1.526, B.1.1.529, and the BA.2 subvariant. Structural analysis reveals that the contact residues of the LY-CoV1404 epitope are highly conserved, except for N439 and N501. Notably, the binding and neutralizing activity of LY-CoV1404 is unaffected by the most common mutations at these positions (N439K and N501Y). The broad and potent neutralization activity and the relatively conserved epitope suggest that LY-CoV1404 has the potential to be an effective therapeutic to treat all known variants.

2.
China Economic Review ; : 101800, 2022.
Article in English | ScienceDirect | ID: covidwho-1797079

ABSTRACT

We consider a model of network interactions where the outcome of a unit depends on the outcomes of the connected units. We determine the key network link, i.e., the network link whose removal results in the largest reduction in the aggregate outcomes, and examine a measure that quantifies the contribution of a network link to the aggregate outcomes. We provide an example examining the spread of Covid-19 in China. Travel restrictions were imposed to limit the spread of infectious diseases. As uniform restrictions can be inefficient and incur unnecessarily high costs, we examine the design of restrictions that target specific travel routes. Our approach may be generalized to multiple countries to guide policies during epidemics ranging from ex ante route-specific travel restrictions to ex post health measures based on travel histories, and from the initial travel restrictions to the phased reopening.

3.
Front Cell Infect Microbiol ; 11: 790422, 2021.
Article in English | MEDLINE | ID: covidwho-1789351

ABSTRACT

Patients with Coronavirus Disease 2019 (COVID-19), due to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection mainly present with respiratory issues and related symptoms, in addition to significantly affected digestive system, especially the intestinal tract. While several studies have shown changes in the intestinal flora of patients with COVID-19, not much information is available on the gut virome of such patients. In this study, we used the viromescan software on the latest gut virome database to analyze the intestinal DNA virome composition of 15 patients with COVID-19 and investigated the characteristic alternations, particularly of the intestinal DNA virome to further explore the influence of COVID-19 on the human gut. The DNA viruses in the gut of patients with COVID-19 were mainly crAss-like phages (35.48%), Myoviridae (20.91%), and Siphoviridae (20.43%) family of viruses. Compared with healthy controls, the gut virome composition of patients with COVID-19 changed significantly, especially the crAss-like phages family, from the first time of hospital admission. A potential correlation is also indicated between the change in virome and bacteriome (like Tectiviridae and Bacteroidaceae). The abundance of the viral and bacterial population was also analyzed through continuous sample collection from the gut of patients hospitalized due to COVID-19. The gut virome is indeed affected by the SARS-CoV-2 infection, and along with gut bacteriome, it may play an important role in the disease progression of COVID-19. These conclusions would be helpful in understanding the gut-related response and contribute to the treatment and prevention strategies of COVID-19.


Subject(s)
COVID-19 , Gastrointestinal Microbiome , DNA , Humans , SARS-CoV-2 , Virome
4.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-331567

ABSTRACT

While humoral immune responses to infection or vaccination with ancestral SARS-CoV-2 have been well-characterized, responses elicited by infection with variants are less understood. Here we characterized the repertoire, epitope specificity, and cross-reactivity of antibodies elicited by Beta and Gamma variant infection compared to ancestral virus. We developed a high-throughput approach to obtain single-cell immunoglobulin sequences and isolate monoclonal antibodies for functional assessment. Spike-, RBD- and NTD-specific antibodies elicited by Beta- or Gamma-infection exhibited a remarkably similar hierarchy of epitope immunodominance for RBD and convergent V gene usage when compared to ancestral virus infection. Additionally, similar public B cell clones were elicited regardless of infecting variant. These convergent responses may account for the broad cross-reactivity and continued efficacy of vaccines based on a single ancestral variant. One Sentence Summary WA1, Beta and Gamma variants of SARS-CoV-2 all elicit antibody responses targeting similar RBD epitopes;public and cross-reactive clones are common.

5.
Science ; : eabn8897, 2022 Mar 24.
Article in English | MEDLINE | ID: covidwho-1759268

ABSTRACT

The rapid spread of the SARS-CoV-2 B.1.1.529 (Omicron) variant and its resistance to neutralization by vaccinee and convalescent sera are driving a search for monoclonal antibodies with potent neutralization. To provide insight into effective neutralization, we determined cryo-EM structures and evaluated receptor-binding domain (RBD) antibodies for their ability to bind and neutralize B.1.1.529. Mutations altered 16% of the B.1.1.529 RBD surface, clustered on a RBD ridge overlapping the ACE2-binding surface and reduced binding of most antibodies. Significant inhibitory activity was retained by select monoclonal antibodies including A19-58.1, B1-182.1, COV2-2196, S2E12, A19-46.1, S309 and LY-CoV1404, which accommodated these changes and neutralized B.1.1.529. We identified combinations of antibodies with synergistic neutralization. The analysis revealed structural mechanisms for maintenance of potent neutralization against emerging variants.

6.
Emerg Microbes Infect ; 11(1): 1010-1013, 2022 Dec.
Article in English | MEDLINE | ID: covidwho-1750052

ABSTRACT

Equine coronavirus (ECoV) was first identified in the USA and has been previously described in several countries. In order to test the presence of ECoV in China, we collected 51 small intestinal samples from donkey foals with diarrhoea from a donkey farm in Shandong Province, China between August 2020 and April 2021. Two samples tested positive for ECoV and full-length genome sequences were successfully obtained using next-generation sequencing, one of which was further confirmed by Sanger sequencing. The two strains shared 100% sequence identity at the scale of whole genome. Bioinformatics analyses further showed that the two Chinese strains represent a novel genetic variant of ECoV and shared the highest sequence identity of 97.05% with the first identified ECoV strain - NC99. In addition, it may be a recombinant, with the recombination region around the NS2 gene. To our knowledge, this is the first documented report of ECoV in China, highlighting its risk to horse/donkey breeding. In addition, its potential risk to public health also warrants further investigation.


Subject(s)
Betacoronavirus 1 , Coronavirus Infections , Horse Diseases , Animals , China/epidemiology , Coronavirus Infections/epidemiology , Coronavirus Infections/veterinary , Diarrhea/veterinary , Equidae , Horse Diseases/epidemiology , Horses , Phylogeny
7.
EuropePMC;
Preprint in English | EuropePMC | ID: ppcovidwho-327388

ABSTRACT

Summary SARS-CoV-2 Omicron is highly transmissible and has substantial resistance to antibody neutralization following immunization with ancestral spike-matched vaccines. It is unclear whether boosting with Omicron-specific vaccines would enhance immunity and protection. Here, nonhuman primates that received mRNA-1273 at weeks 0 and 4 were boosted at week 41 with mRNA-1273 or mRNA-Omicron. Neutralizing antibody titers against D614G were 4760 and 270 reciprocal ID 50 at week 6 (peak) and week 41 (pre-boost), respectively, and 320 and 110 for Omicron. Two weeks after boost, titers against D614G and Omicron increased to 5360 and 2980, respectively, for mRNA-1273 and 2670 and 1930 for mRNA-Omicron. Following either boost, 70-80% of spike-specific B cells were cross-reactive against both WA1 and Omicron. Significant and equivalent control of virus replication in lower airways was observed following either boost. Therefore, an Omicron boost may not provide greater immunity or protection compared to a boost with the current mRNA-1273 vaccine.

8.
EuropePMC;
Preprint in English | EuropePMC | ID: ppcovidwho-327132

ABSTRACT

Immunization with SARS-CoV-2 spike elicits diverse antibodies, but can any of these neutralize broadly? Here, we report the isolation and characterization of antibody WS6, from a mouse immunized with mRNA encoding the SARS-CoV-2 spike. WS6 bound diverse beta-coronavirus spikes and neutralized SARS-CoV-2 variants, SARS-CoV, and related sarbecoviruses. Epitope mapping revealed WS6 to target a region in the S2 subunit, which was conserved among SARS-CoV-2, MERS-CoV, and hCoV-OC43. The crystal structure at 2-angstrom resolution of WS6 with its S2 epitope revealed recognition to center on a conserved helix, which was occluded in both prefusion and post-fusion spike conformations. Structural and neutralization analyses indicated WS6 to neutralize by inhibiting fusion, post-viral attachment. Comparison of WS6 to other antibodies recently identified from convalescent donors or mice immunized with diverse spikes indicated a stem-helical supersite - centered on hydrophobic residues Phe1148, Leu1152, Tyr1155, and Phe1156 - to be a promising target for vaccine design.

9.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-318615

ABSTRACT

Background: To explore the changes in lymphocyte subsets and cytokine profiles in patients with coronavirus disease 2019 (COVID-19) and their relationship with disease severity. Methods: : This study included 228 patients with COVID-19 who were treated at Chongqing University Three Gorges Hospital from January 1, 2020 to February 20, 2020. The characteristics of lymphocyte subsets and cytokine profiles of severe and mild COVID-19 patients were compared. Of the 228 patients enrolled, 48 were severe patients and 180 were mild patients. Results: : Lymphocyte counts, absolute number of total T lymphocytes, CD 4+ T cells, CD 8+ T cells, and total B lymphocytes were significantly lower in severe patients (0.8×10 9 /L, 424.5×10 6 /L, 266×10 6 /L, 145.5×10 6 /L, 109.5×10 6 /L, respectively) than in mild patients (1.2×10 9 /L, 721×10 6 /L, 439.5×10 6 /L, 281.5×10 6 /L, 135×10 6 /L, respectively). A multivariate logistic regression analysis showed that age, C-reactive protein (CRP) and the neutrophil-to-lymphocyte ratio (NLR) were independent risk factors for developing into severe condition. The lymphocyte subsets decreased and cytokine profiles increased more significantly in severe patients than in mild patients. Conclusions: : CRP, NLR, and age may serve as powerful factors for early identification of severe patients.

10.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-308278

ABSTRACT

We consider a model of network interactions where the outcome of a unit depends on the outcomes of the connected units. We determine the key network link, i.e., the network link whose removal results in the largest reduction in the aggregate outcomes, and provide a measure that quantifies the contribution of a network link to the aggregate outcomes, which complements the intercentrality measure of the key network node proposed by Ballester, Calv ́o-Armengol, and Zenou (2006). We provide an example examining the spread of Covid-19 in China. Travel restrictions were imposed to limit the spread of infectious diseases. As uniform restrictions can be inefficient and incur unnecessarily high costs, we examine the design of restrictions that target specific travel routes. Our approach may be generalized to multiple countries to guide policies during epidemics ranging from ex ante route-specific travel restrictions to ex post health measures based on travel histories, and from the initial travel restrictions to the phased reopening.

11.
mBio ; : e0322721, 2022 Feb 15.
Article in English | MEDLINE | ID: covidwho-1685493

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) harbor mutations in the spike (S) glycoprotein that confer more efficient transmission and dampen the efficacy of COVID-19 vaccines and antibody therapies. S mediates virus entry and is the primary target for antibody responses, with structural studies of soluble S variants revealing an increased propensity toward conformations accessible to the human angiotensin-converting enzyme 2 (hACE2) receptor. However, real-time observations of conformational dynamics that govern the structural equilibriums of the S variants have been lacking. Here, we report single-molecule Förster resonance energy transfer (smFRET) studies of critical mutations observed in VOCs, including D614G and E484K, in the context of virus particles. Investigated variants predominately occupied more open hACE2-accessible conformations, agreeing with previous structures of soluble trimers. Additionally, these S variants exhibited slower transitions in hACE2-accessible/bound states. Our finding of increased S kinetic stability in the open conformation provides a new perspective on SARS-CoV-2 adaptation to the human population. IMPORTANCE SARS-CoV-2 surface S glycoprotein-the target of antibodies and vaccines-is responsible for binding to the cellular receptor hACE2. The interactions between S and hACE2 trigger structural rearrangements of S from closed to open conformations prerequisite for virus entry. Under the selection pressure imposed by adaptation to the human host and increasing vaccinations and convalescent patients, SARS-CoV-2 is evolving and has adopted numerous mutations on S variants. These promote virus spreading and immune evasion, partially by increasing the propensity of S to adopt receptor-binding competent open conformations. Here, we determined a time dimension, using smFRET to delineate the temporal prevalence of distinct structures of S in the context of virus particles. We present the first experimental evidence of decelerated transition dynamics from the open state, revealing increased stability of S open conformations to be part of the SARS-CoV-2 adaption strategies.

12.
Nat Hum Behav ; 6(3): 311, 2022 03.
Article in English | MEDLINE | ID: covidwho-1671567

Subject(s)
COVID-19 , Humans , SARS-CoV-2
13.
Front Pharmacol ; 12: 759587, 2021.
Article in English | MEDLINE | ID: covidwho-1662607

ABSTRACT

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of coronavirus disease 2019 (COVID-19), which has resulted in a global pandemic. Methodology: We used a two-step polymerase chain reaction to detect the ACE genotype and ELISA kits to detect the cytokine factor. We also used proteomics to identify the immune pathway related to the ACE protein expression. Result: In this study, we found that the angiotensin-converting enzyme (ACE) deletion polymorphism was associated with the susceptibility to COVID-19 in a risk-dependent manner among the Chinese population. D/D genotype distributions were higher in the COVID-19 disease group than in the control group (D/D odds ratio is 3.87 for mild (p value < 0.0001), 2.59 for moderate (p value = 0.0002), and 4.05 for severe symptoms (p value < 0.0001), logic regression analysis. Moreover, genotype-specific cytokine storms and immune responses were found enriched in patients with the ACE deletion polymorphism, suggesting the contribution to the susceptibility to COVID-19. Finally, we identified the immune pathway such as the complement system related to the ACE protein expression of patients by lung and plasma proteomics. Conclusion: Our results demonstrated that it is very important to consider gene polymorphisms in the population to discover a host-based COVID-19 vaccine and drug design for preventive and precision medicine.

14.
Front Public Health ; 9: 767004, 2021.
Article in English | MEDLINE | ID: covidwho-1598043

ABSTRACT

Background: The outbreak of the COVID-19 pandemic has had a profound influence on the mental health and well-being of individuals across the globe. Emotional competence, defined as one's ability to recognize, understand, and manage their emotions, has been found linked with mental health problems (e.g., depression and anxiety) in previous studies. However, there is limited knowledge about the direction of the association between these factors among populations exposed to COVID-19. This study examined the possible mediation relationships between depression, anxiety, emotional competence, and COVID-19 exposure among Chinese adolescents. Methods: Responses from 7,958 Chinese adolescents who had previously taken part in a two-wave study before (December 23, 2019-January 13, 2020) and during COVID-19 (June 16, 2020-July 8, 2020) were analyzed (51.67% males, mean age = 11.74, SD = 2.15). Structural equation modeling with three covariates (i.e., age, gender, and ethnicity) was used to test the longitudinal mediation relationships between COVID-19 exposure and depression, anxiety via emotional competence. Results: Results indicated that the prevalence of depression (38.67 to 36.74%) and anxiety (13.02 to 12.77%) decreased from Time 1 to Time 2. The T2 emotional competence significantly mediated the relationship between T2 COVID-19 exposure and T2 anxiety (indirect effect [95% CI] = 0.011 [0.004-0.019], p < 0.05). T2 emotional competence also significantly mediated the relationship between T2 COVID-19 exposure and T2 depression (indirect effect [95% CI] = 0.013 [0.005-0.022], p < 0.05). The results indicated that T2 emotional competence had a significant and negative influence on T2 anxiety (ß = -0.266, SE = 0.005, p < 0.001), and T2 depression (ß = -0.326, SE = 0.029, p < 0.001). Conclusions: This longitudinal research study demonstrated the crucial role of emotional competence in influencing the severity of long-term mental health problems, and suggested that emotional competence interventions can be conducted to improve mental well-being among Chinese adolescents exposed to COVID-19.


Subject(s)
COVID-19 , Adolescent , Anxiety Disorders , Child , China/epidemiology , Depression/epidemiology , Emotions , Female , Humans , Male , Pandemics , SARS-CoV-2
15.
Cell ; 185(1): 113-130.e15, 2022 01 06.
Article in English | MEDLINE | ID: covidwho-1588150

ABSTRACT

mRNA-1273 vaccine efficacy against SARS-CoV-2 Delta wanes over time; however, there are limited data on the impact of durability of immune responses on protection. Here, we immunized rhesus macaques and assessed immune responses over 1 year in blood and upper and lower airways. Serum neutralizing titers to Delta were 280 and 34 reciprocal ID50 at weeks 6 (peak) and 48 (challenge), respectively. Antibody-binding titers also decreased in bronchoalveolar lavage (BAL). Four days after Delta challenge, the virus was unculturable in BAL, and subgenomic RNA declined by ∼3-log10 compared with control animals. In nasal swabs, sgRNA was reduced by 1-log10, and the virus remained culturable. Anamnestic antibodies (590-fold increased titer) but not T cell responses were detected in BAL by day 4 post-challenge. mRNA-1273-mediated protection in the lungs is durable but delayed and potentially dependent on anamnestic antibody responses. Rapid and sustained protection in upper and lower airways may eventually require a boost.

16.
Frontiers in cellular and infection microbiology ; 11, 2021.
Article in English | EuropePMC | ID: covidwho-1564449

ABSTRACT

Patients with Coronavirus Disease 2019 (COVID-19), due to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection mainly present with respiratory issues and related symptoms, in addition to significantly affected digestive system, especially the intestinal tract. While several studies have shown changes in the intestinal flora of patients with COVID-19, not much information is available on the gut virome of such patients. In this study, we used the viromescan software on the latest gut virome database to analyze the intestinal DNA virome composition of 15 patients with COVID-19 and investigated the characteristic alternations, particularly of the intestinal DNA virome to further explore the influence of COVID-19 on the human gut. The DNA viruses in the gut of patients with COVID-19 were mainly crAss-like phages (35.48%), Myoviridae (20.91%), and Siphoviridae (20.43%) family of viruses. Compared with healthy controls, the gut virome composition of patients with COVID-19 changed significantly, especially the crAss-like phages family, from the first time of hospital admission. A potential correlation is also indicated between the change in virome and bacteriome (like Tectiviridae and Bacteroidaceae). The abundance of the viral and bacterial population was also analyzed through continuous sample collection from the gut of patients hospitalized due to COVID-19. The gut virome is indeed affected by the SARS-CoV-2 infection, and along with gut bacteriome, it may play an important role in the disease progression of COVID-19. These conclusions would be helpful in understanding the gut-related response and contribute to the treatment and prevention strategies of COVID-19.

17.
Science ; 373(6556)2021 Aug 13.
Article in English | MEDLINE | ID: covidwho-1559379

ABSTRACT

The emergence of highly transmissible SARS-CoV-2 variants of concern (VOCs) that are resistant to therapeutic antibodies highlights the need for continuing discovery of broadly reactive antibodies. We identified four receptor binding domain-targeting antibodies from three early-outbreak convalescent donors with potent neutralizing activity against 23 variants, including the B.1.1.7, B.1.351, P.1, B.1.429, B.1.526, and B.1.617 VOCs. Two antibodies are ultrapotent, with subnanomolar neutralization titers [half-maximal inhibitory concentration (IC50) 0.3 to 11.1 nanograms per milliliter; IC80 1.5 to 34.5 nanograms per milliliter). We define the structural and functional determinants of binding for all four VOC-targeting antibodies and show that combinations of two antibodies decrease the in vitro generation of escape mutants, suggesting their potential in mitigating resistance development.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/immunology , Angiotensin-Converting Enzyme 2/antagonists & inhibitors , Angiotensin-Converting Enzyme 2/metabolism , Antibodies, Neutralizing/chemistry , Antibodies, Neutralizing/metabolism , Antibodies, Viral/chemistry , Antibodies, Viral/metabolism , Antibody Affinity , Antigen-Antibody Reactions , COVID-19/virology , Humans , Immune Evasion , Immunoglobulin Fab Fragments/immunology , Immunoglobulin Fab Fragments/metabolism , Mutation , Neutralization Tests , Protein Domains , Receptors, Coronavirus/antagonists & inhibitors , Receptors, Coronavirus/metabolism , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism
18.
2021.
Preprint in English | Other preprints | ID: ppcovidwho-296309

ABSTRACT

LY-CoV1404 (also known as bebtelovimab) is a highly potent, neutralizing, SARS-CoV-2 spike glycoprotein receptor binding domain (RBD)-specific antibody identified from a convalescent COVID-19 patient sample, obtained approximately 60 days after symptom onset. In pseudovirus studies, LY-CoV1404 retains potent neutralizing activity against numerous variants including B.1.617.2, B.1.1.7, B.1.351, B.1.427/B.1.429, P.1, and B.1.526, binding to these variants in the presence of their underlying RBD mutations (which include K417N, L452R, E484K, and N501Y). LY-CoV1404 also neutralizes multiple isolates of the authentic SARS-CoV-2 virus in two different assays. The RBD positions comprising the LY-CoV1404 epitope are highly conserved, with the exception of N439 and N501;notably the binding and neutralizing activity of LY-CoV1404 is unaffected by the most common mutations at these positions (N439K and N501Y). New variant-resistant treatments such as LY-CoV1404 are desperately needed, given that some of the existing therapeutic antibodies are less effective or ineffective against certain variants and the impact of variants on vaccine efficacy is still poorly understood. The breadth of variant binding, potent neutralizing activity and the relatively conserved epitope suggest that LY-CoV1404 is one in a panel of well-characterized, clinically developable antibodies that could be deployed as potentially long-term solutions to address current and emerging variants. In Brief LY-CoV1404 is a potent SARS-CoV-2-binding antibody that neutralizes all known variants of concern and whose epitope is rarely mutated. Highlights LY-CoV1404 potently neutralizes SARS-CoV-2 authentic virus and all known variants of concern including the B.1.617.2 (Delta) variant No loss of potency against currently circulating variants Binding epitope on RBD of SARS-CoV-2 is rarely mutated in GISAID database Breadth of neutralizing activity and potency supports clinical development

19.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-293958

ABSTRACT

This paper models the local and cross-city transmissions of the novel coronavirus in China between January 19 and February 29 in 2020. We examine the role of various socioeconomic mediating factors, including public health measures that encourage social distancing in local communities. Weather characteristics two weeks ago are used as instrumental variables for causal inference. Stringent quarantine, city lockdown, and local public health measures imposed since late January significantly decreased the virus transmission rate.The virus spread was contained by the middle of February. Population outflow from the outbreak source region posed a higher risk to the destination regions than other factors including geographic proximity and similarity in economic conditions. We quantify the effects of different public health measures in reducing the number of infections through counterfactual analyses. Over 1.4 million infections and 56,000 deaths could have been avoided as a result of the national and provincial public health measures imposed in late January in China.

20.
PubMed; 2020.
Preprint in English | PubMed | ID: ppcovidwho-292815

ABSTRACT

Biotin-labeled molecular probes, comprising specific regions of the SARS-CoV-2 spike, would be helpful in the isolation and characterization of antibodies targeting this recently emerged pathogen. To develop such probes, we designed constructs incorporating an N-terminal purification tag, a site-specific protease-cleavage site, the probe region of interest, and a C-terminal sequence targeted by biotin ligase. Probe regions included full-length spike ectodomain as well as various subregions, and we also designed mutants to eliminate recognition of the ACE2 receptor. Yields of biotin-labeled probes from transient transfection ranged from ~0.5 mg/L for the complete ectodomain to >5 mg/L for several subregions. Probes were characterized for antigenicity and ACE2 recognition, and the structure of the spike ectodomain probe was determined by cryo-electron microscopy. We also characterized antibody-binding specificities and cell-sorting capabilities of the biotinylated probes. Altogether, structure-based design coupled to efficient purification and biotinylation processes can thus enable streamlined development of SARS-CoV-2 spike-ectodomain probes. Funding: Support for this work was provided by the Intramural Research Program of the Vaccine Research Center, National Institute of Allergy and Infectious Diseases (NIAID). Support for this work was also provided by COVID-19 Fast Grants, the Jack Ma Foundation, the Self Graduate Fellowship Program, and NIH grants DP5OD023118, R21AI143407, and R21AI144408. Some of this work was performed at the Columbia University Cryo-EM Center at the Zuckerman Institute, and some at the Simons Electron Microscopy Center (SEMC) and National Center for Cryo-EM Access and Training (NCCAT) located at the New York Structural Biology Center, supported by grants from the Simons Foundation (SF349247), NYSTAR, and the NIH National Institute of General Medical Sciences (GM103310). Conflict of Interest: The authors declare that they have no conflict of interest. Ethical Approval: Peripheral blood mononuclear cells (PBMCs) for B cell sorting were obtained from a convalescent SARS-CoV-2 patient (collected 75 days post symptom onset under an IRB approved clinical trial protocol, VRC 200 - ClinicalTrials.gov Identifier: NCT00067054) and a healthy control donor from the NIH blood bank pre-SARS-CoV-2 pandemic.

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