Anti-Zika Virus (ZIKV) E Protein [Clone ZV67] — Purified in vivo GOLD™ Functional Grade

The ZV-67 monoclonal antibody is used in in vivo mouse studies primarily for passive immunotherapy to test protective efficacy against Zika virus (ZIKV) infection. This antibody targets the lateral ridge of domain III (DIII-LR) of the Zika virus envelope protein and has demonstrated significant protective activity in multiple experimental models.

Passive Transfer Protection Studies

The most common application of ZV-67 in mouse studies involves passive transfer experiments where the antibody is administered to mice before viral challenge. In these studies, mice receive 250 ?g of ZV-67 monoclonal antibody via intraperitoneal injection one day before being infected with ZIKV. This approach allows researchers to evaluate the direct protective capacity of the antibody without requiring the mouse's immune system to generate its own antibodies.

Protection Against Multiple ZIKV Strains

ZV-67 has shown protective efficacy against different ZIKV strains in various mouse models. The antibody provided significant protection against ZIKV Dakar 41519 infection in young mice and demonstrated effectiveness against the ZIKV Asian strain GZ01 infection in pregnant mice. Additionally, controlled studies using C57BL/6 mice showed that ZV-67 protected against lethal ZIKV Dakar 41519 challenge, with treated mice showing significantly better survival compared to control groups receiving non-specific antibodies.

Experimental Protocol and Monitoring

In typical in vivo studies, researchers use a standardized protocol where 4-5 week-old C57BL/6 mice are first treated with 2 mg of anti-Ifnar1 monoclonal antibody to suppress interferon signaling, making them more susceptible to ZIKV infection. The mice then receive 250 ?g of ZV-67 before being challenged with 10^5 focus-forming units (FFU) of ZIKV via subcutaneous inoculation. Researchers monitor the mice by measuring daily body weights and collecting serum samples on day 3 post-infection to analyze viral RNA levels using quantitative RT-PCR.

Research Applications

Beyond protection studies, ZV-67 serves as a valuable research tool for studying ZIKV pathogenesis and immune responses. The antibody is used in various laboratory techniques including ELISA, Western blot, flow cytometry, and neutralization assays such as focus reduction neutralization tests (FRNT) and plaque reduction neutralization tests (PRNT). Its specificity for ZIKV without cross-reactivity to related flaviviruses like dengue or Japanese encephalitis virus makes it particularly useful for distinguishing ZIKV-specific immune responses.

This mouse monoclonal antibody has become an important tool for evaluating potential therapeutic interventions and understanding the mechanisms of antibody-mediated protection against Zika virus infection in preclinical models.

Clone ZV-67 is a monoclonal antibody that specifically binds to the Zika virus envelope (E) protein, and its proper storage is critical for maintaining stability and effectiveness.

Short-term Storage

For immediate use, clone ZV-67 should be stored at 2-8°C (36-46°F) for up to one month. The antibody is provided sterile as received and can maintain its integrity within this refrigerated temperature range for short-term applications.

Long-term Storage

For extended storage periods, clone ZV-67 requires ?-70°C (-94°F or below). When storing long-term, it's essential to aseptically aliquot the antibody into working volumes without diluting the original concentration. This approach helps maintain the antibody's functional properties over extended periods.

Critical Storage Considerations

Avoid repeated freeze-thaw cycles as this can significantly compromise the antibody's structure and binding capacity. The antibody is formulated in phosphate buffered saline (PBS) at pH 7.2-7.4 without carrier proteins, potassium, calcium, or preservatives, which makes it particularly sensitive to temperature fluctuations.

Due to the inherent biochemical properties of antibodies, precipitation may occur over time during storage. If this happens, the precipitate can be removed through aseptic centrifugation and/or filtration without affecting the antibody's functionality.

The antibody maintains high purity standards with ?95% monomer content and endotoxin levels below 1.0 EU/µg, making proper temperature control essential to preserve these quality characteristics.

Commonly Used Antibodies and Proteins with ZV-67 in Zika Virus Research

ZV-67 is a well-characterized mouse monoclonal antibody (mAb) that binds the envelope (E) protein of Zika virus (ZIKV), particularly targeting domain III (DIII), and demonstrates potent neutralizing activity against both African and Asian ZIKV strains. In the literature, ZV-67 is frequently used alongside other antibodies and proteins for comparative studies in neutralization, epitope mapping, and structural analyses.

Key Antibodies Used with ZV-67

Proteins Commonly Used with ZV-67

• ZIKV E Protein (Domains I, II, III): The primary target for ZV-67, especially DIII, but also engaging epitopes in DI and DII in some studies.
• ZIKV Non-Structural Proteins: Sometimes included in epitope discovery studies, though ZV-67 itself targets the E protein.
• Biotin-labeled DIII: Used in biolayer interferometry (BLI) to study epitope competition between ZV-67 and related antibodies.

Experimental Contexts

• Epitope mapping and competition: ZV-67 is compared with ZV-2, ZV-48, ZV-54, and ZV-64 using BLI to identify overlapping and distinct epitopes on the ZIKV E protein.
• Neutralization assays: ZV-67 is tested alongside other mAbs (e.g., ZV-54, CHK-166) in mouse models to evaluate protection against lethal ZIKV challenge.
• Structural studies: Ribbon diagrams and crystal structures of ZV-67–DIII complexes are compared with those of ZV-2, ZV-48, and ZV-64 to elucidate antibody-binding sites.
• Molecular docking: ZV-67 and Z3L1 are used to dock with novel E protein epitopes from Indian ZIKV strains, revealing conserved and strain-specific interactions.
• Serological controls: In ELISA and neutralization assays, ZV-67 is used alongside 4G2 (for DI/II) and MERS-4 (negative control) to ensure specificity.

Summary Table: Common Antibodies and Proteins Used with ZV-67

Takeaway

ZV-67 is most often used with a panel of ZIKV DIII-specific mouse mAbs (ZV-2, ZV-48, ZV-54, ZV-64) for neutralization, structural, and epitope competition studies. It is also paired with control antibodies like 4G2 and MERS-4 in serological assays, and with the human mAb Z3L1 in epitope mapping of emerging ZIKV strains. The primary protein target is the ZIKV E protein, especially DIII, but work also extends to DI and DII in some contexts.

ZV-67 is a Zika virus (ZIKV)-specific monoclonal antibody that has been extensively studied for its neutralizing properties and structural characteristics. The key findings from scientific literature reveal several important insights about this antibody clone and its therapeutic potential.

Neutralizing Activity and Strain Specificity

ZV-67 demonstrates potent neutralizing activity against multiple ZIKV strains, showing broad-spectrum effectiveness compared to other antibodies in the same panel. Unlike some related antibodies such as ZV-48 and ZV-64, which showed reduced inhibitory activity against American and African ZIKV strains, ZV-67 maintained efficient neutralization across different viral strains. This broad neutralizing capacity makes it particularly valuable for potential therapeutic applications.

Epitope Recognition and Structural Binding

The antibody recognizes a highly specific epitope on the ZIKV envelope protein domain III (DIII) known as the lateral ridge (LR) epitope. This epitope is created by four discrete secondary structure elements: the A-strand, B-C loop, D-E loop, and F-G loop. ZV-67 makes contact with a total of 21 residues within this epitope region, with only one amino acid difference observed between different ZIKV immunizing strains (E^E393^ in H/PF/2013 versus D^E393^ in MR-766).

Remarkably, the LR epitope recognized by ZV-67 shares significant structural homology with epitopes targeted by other flavivirus antibodies. It shares 13 contact positions with the potently neutralizing E16 antibody that targets West Nile virus, and 10 contact positions with the DV1-E106 antibody. This cross-reactivity pattern suggests evolutionary conservation of critical neutralizing sites across flaviviruses.

Protective Efficacy in Animal Models

In vivo passive transfer studies demonstrated that ZV-67 provides protective activity against lethal ZIKV challenge in mouse models. Specifically, the antibody showed protection against an African ZIKV strain, indicating its potential as a therapeutic intervention for ZIKV infection. This protective capacity was shared with ZV-54, another antibody with similar functional properties.

Use in Vaccine Development and Epitope Mapping

ZV-67 has proven valuable as a tool for vaccine development and epitope mapping studies. Researchers have used this antibody in phage display selections to identify resurfaced ZIKV EDIII variants that maintain conformationally relevant epitopes while altering other surface characteristics. The antibody serves as a positive selection target to ensure that engineered immunogens retain critical neutralizing epitopes.

During these studies, ZV-67 was used to validate that engineered variants maintained an intact lateral ridge epitope, while other regions of the protein (ABDE sheet and CC' loop) were successfully altered. This application demonstrates the antibody's utility in structure-guided vaccine design approaches.

Molecular Characteristics and Clonal Relationships

Sequence analysis revealed that ZV-67 shares highly similar variable light (V_L) and variable heavy (V_H) chain sequences with ZV-54, another functionally related monoclonal antibody. This similarity explains their comparable neutralizing properties and protective efficacy. The close molecular relationship between these antibodies suggests they may have arisen from similar B-cell responses or represent optimized variants targeting the same critical viral epitope.

These findings collectively establish ZV-67 as a lead candidate for ZIKV therapeutics and a valuable research tool for understanding flavivirus neutralization mechanisms and vaccine design.