Recombinant Proteins

Virus-like particles (VLPs) are small nanoparticles composed of the structural proteins of a virus, specifically the shell or capsid proteins. These proteins self-assemble into virus-like structures, mimicking the organization and morphology of the actual virus. However, VLPs are devoid of the viral infectious genomes, rendering them non-infectious and relatively safe for use in various applications, including vaccine development and gene therapy.

Mechanism of expression for VLP-displayed antigens.

Why use VLP-displayed antigens?

Virus-Like Particles (VLPs) are good at boosting the immune response. This makes them useful for immunization, especially with antigen targets that are usually present in low amounts or don’t trigger a strong immune reaction by themselves. They also allow for soluble expression of multipass transmembrane proteins in their natural configuration.

Applications

→ Blood sample determination: ELISA

→ In vivo pharmacokinetic analysis

→ Antibody Immunization, Screening, Functional Characterization

→ CMC method development

→ Affinity determination: ELISA, SPR

Features

→ Site-Specific Biotinylation

→ VLP framework optimized to each antigen

→ Proven biological activity via ELISA and SPR

→ Mammalian cell expression system for natural folding and glycosylation

→ Boosted immunogenicity for antibody drug discovery

Product Validation Data

Claudin 18.2 VLP

In 2018, KACTUS became the first company globally to express full-length wild-type Claudin 18.2, a multi-transmembrane protein with significant implications for gastric and esophageal adenocarcinomas. Despite its potential, technical challenges in generating high-quality Claudin 18.2 antigen have previously hindered antibody isolation and production quality control. Leveraging our SAMS™ protein engineering and expression platform, we have successfully produced full-length Claudin 18.2 antigens on VLPs. Our robust product performance testing demonstrates the functional integrity and bioactivity of our VLP expression platform.

Figure 1. This graph illustrates the intensity distribution of Claudin 18.2 VLPs as measured by dynamic light scattering (DLS). The peak centered at approximately 150 nm indicates a homogeneous population of VLPs, reflecting the consistent size and quality of the particles.

Figure 2. The chromatogram shows the high-performance liquid chromatography (HPLC) profile of Claudin 18.2 VLPs, with a prominent peak observed at 4.909 minutes, indicating the retention time of the Claudin 18.2 protein. The sharp and well-defined peak suggests a high purity and consistency of the VLP preparation.

Figure 3. ELISA assay shows the binding activity of Claudin 18.2 VLPs to anti-Claudin 18.2 antibodies across three different production batches. The consistent curves for Batches 1, 2, and 3 highlight the reproducibility and stability of the VLP preparations. The half-maximal effective concentration (EC50) is determined to be 0.1 nM, indicating high affinity and robust interaction between Claudin 18.2 VLPs and the specific antibodies.

Figure 4. Biolayer Interferometry (BLI) data demonstrates the binding kinetics of Claudin 18.2 VLPs to streptavidin-labeled probes. The analysis reveals a high binding affinity with a dissociation constant (KD) of approximately 5.73E-10 M, indicating strong and stable interaction.

Figure 5. Flow cytometry data demonstrates the binding specificity and quantification of Claudin 18.2 displayed on VLPs. The top panels show the results from HEK293 control cells, while the bottom panels show the results from anti-Cld18.2 CAR-expressing HEK293 cells. The presence of Cld18.2 VLPs results in a significant shift in the fluorescence signal in anti-Cld18.2 CAR-expressing cells, indicating successful detection and quantification of Cld18.2 on VLPs. This validates the high-quality expression and bioactivity of the Claudin 18.2 VLPs.

Figure 6. SPR data showing the binding interactions of Biotinylated Claudin 18.2 VLPs with streptavidin immobilized on a CM5 chip. Results show a dissociation constant (KD) of 1.638E-11 M. This indicates a very strong and stable binding affinity of the Claudin 18.2 VLP.

VLP Products

Nanodiscs are membrane-like structures composed of phospholipids and membrane scaffold molecules, which are typically synthetic copolymers or membrane scaffold proteins. We use a proprietary amphipathic copolymer to enable membrane protein stabilization & solubilization without the use of detergents.

Schematic diagram of nanodisc displaying a multi-transmembrane protein

Why use nanodisc-displayed antigens?

Using Nanodisc technology to display multitransmembrane proteins not only better maintains their natural conformation but also addresses the issue of soluble preparation of these proteins. Moreover, because nanodiscs can be extracted without detergents, they offer a superior alternative for multitransmembrane proteins that are sensitive to maintaining activity with detergents.

Applications

→ Yeast display screening
→ In vitro functional assays
→ CAR expression testing
→ PK/PD Studies
→ Analytical Tests, ELISA, SPR, BLI

Features

→ Native structure and conformation
→ Detergent free extraction for reliable performance
→ Good water solubility
→ Proven biological activity via ELISA and SPR
→ Mammalian cell expression system for natural folding and glycosylation

Product Validation Data

GPRC5D Nanodisc

GPRC5D is an important multitransmembrane protein with significant therapeutic and research potential. To ensure the most accurate representation of its natural conformation and activity, we’ve developed a GPRC5D Nanodisc with detergent-free extraction. This innovative approach not only maintains the native structure of GPRC5D but also overcomes the challenges associated with the soluble preparation of multi-transmembrane proteins. We’ve performed extensive product quality testing to demonstrate the efficacy and quality of our Nanodisc-based antigens.

Additionally, to further support the development of antibody drugs (such as bispecific antibodies), we’ve developed biotinylated Nanodisc-displayed multitransmembrane proteins. These proteins exhibit high binding activity and can be applied to various research stages, including ELISA method development and pharmacokinetic analysis.

Figure 7. Human GPRC5D Nanodisc with His Tag was immobilized at 2 µg/ml (100 µl/well) on the plate. The dose-response curve for the Anti-GPRC5D Antibody with hFc Tag was generated, with an EC50 of 4.9 ng/ml as determined by ELISA.

Figure 8. Human CD3E&CD3D, with a His Tag, was immobilized on the plate at a concentration of 2 μg/mL (100 μL/well). Serial dilutions of the Anti-Human CD3×GPRC5D bispecific antibody, with an hFc Tag, were then added, followed by the addition of biotinylated Human GPRC5D Nanodisc, with a His Tag, at 5 μg/mL. Detection was performed using HRP-conjugated streptavidin, and the EC50 was determined to be 0.28 μg/mL by ELISA.

Figure 9. Biotinylated Human GPRC5D Nanodisc, with a His Tag, was loaded onto an SA-Biosensor. This setup was used to determine the binding affinity to the Anti-GPRC5D Antibody, which was measured to be 1.16 nM using a BLI assay.

Figure 10. Human GPRC5D Nanodisc with His Tag was captured on a CM5 Chip via an anti-His antibody. It was found to bind the Anti-GPRC5D Antibody with an affinity constant of 1.47 nM, as determined by SPR assay (Biacore T200).

 Nanodisc Products

KACTUS VLP-Displayed Antigens FAQs

Product Features

High Protein Affinity
We verified the activity of our heterodimers, such as CD3E/G-His Tag and CD3E/D-His Tag, via ELISA and SPR.

Equal Expression of Subunits
A proprietary design, expression, and purification system ensures the two subunits of the heterodimers are expressed in equal proportions with greater than 95% purity, verfied by SDS-PAGE and SEC-HPLC.

Stability
Our CD3 proteins have high batch-to-batch consistency and long-term stability as verified by ELISA.

CD3 Proteins Overview

About CD3 Proteins

CD3 (Cluster of differentiation 3) is one of the proteins expressed on the surface of T cells. It consists of six peptide chains, one γ chain, one δ chain, two ε chains and two ζ chains. These six peptide chains are all transmembrane, and the transmembrane region is connected with TCR through a salt bridge to form a tight TCR-CD3 complex, which jointly participates in the recognition and signal transduction of T cells to antigens. The activation signal generated by TCR recognition of antigen is transduced into T cells by the ITAM sequence in the cytoplasmic region of CD3, and then a series of immune responses are initiated.

Applications

Antibody Discovery

Immunization

Antibody Screening

Functional characterization

Affinity Determination (ELISA, SPR, BLI)

TCR/CD3 signaling (Louis-Dit-Sully C et. al, 2012）

Product Performance Validation

High Binding Affinity by SPR

Human CD3E/CD3D His tag, captured on CM5 chip via anti-His antibody, binds OKT3 mFc Tag with an affinity constant of 0.36nM as determined in SPR assay (Biacore T200).

Batch-to-Batch Consistency

Immobilized Human CD3E/CD3G, hFc Tag, at 1μg/mL (100μL/well). Dose-response curve for anti-CD3E/CD3G antibody (OKT3, mFc Tag). The EC50s are 20.7, 21.0 and 20.ong/mL respectively as determined by ELISA.

Temperature Stability

Immobilized Human CD3E/CD3G, hFc Tag, at 1μg/mL (100μL/well). Dose-response curve for anti-CD3E/CD3G antibody (OKT3, mFc Tag). The EC50s are 20.7, 19.5, 20.0 and 20.7ng/mL respectively, as determined by ELISA

High Purity by SEC-HPLC

The purity of Cynomolgus CD3E is greater than 95% as determined by SEC-HPLC.

High Purity by Tris-Bis PAGE

Cynomolgus CD3E/CD3D on Tris-Bis PAGE under reduced conditions. The purity is greater than 95%.

Browse CD3 Proteins

Browse our popular CD3 proteins here or click below to view the full catalog selection:

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Background

FGFR (Fibroblast Growth Factor Receptors), generally includes four types of FGFR1-4 (also known as CD331-334). FGFR is a branch of the tyrosinase receptor family, which is a type I transmembrane protein usually functioning as a dimer. Among them, the extracellular domain contains three Ig-like regions D1 (IgI), D2 (IgII), and D3 (IgIII), of which D1 and the Acidic box form an autoinhibitory region, and D2 and D3 are responsible for ligand binding (D2 and cell surface Heparan sulfate is combined, D3 has two forms of IIIb and IIIc due to alternative splicing, and only one isoform of IIIc is currently found in FGFR4). According to the number of Ig-like domains, FGFR can be divided into two forms, one is α-type, which contains three regions of IgI, IgII, and IgIII; the other is β-type, which only contains IgII and IgIII.

Figure 1. FGFR structure (Babina IS, et al., 2017)

The ligand of FGFR is FGF. There are 18 kinds of human FGFs that have been discovered. Except for FGF19, FGF21, and FGF23, which are endocrine, the rest are paracrine. Among them, FGF7 can only bind to type IIIb FGFR2. The combination of FGFR and FGF mediates the activation and transmission of signaling pathways such as RAS-RAF-MAPK, PI3K-AKT, JAK-STAT and PLCγ, and participates in important physiological functions such as cell growth, differentiation, migration, neovascularization, regulation of organ development and wound healing. FGFR gene mutations are commonly found in solid tumors such as lung cancer, liver cancer, intrahepatic cholangiocarcinoma, breast cancer, gastric cancer, uterine cancer, and bladder cancer, and there are differences in the types and frequencies of FGFR mutations in different cancer types.

Figure 2. FGF-FGFR-mediated signaling pathway (Mason I, 2007)

KACTUS has deeply analyzed the structural differences of FGFR family proteins and successfully prepared various types of FGFR recombinant proteins, covering α and β isoforms, as well as different forms such as IIIb and IIIc, supporting the differentiated research of FGFR-targeted drugs.

Product Performance Validation

Human FGFR2 beta (IIIb) Protein (FGR-HM1BB)

Figure 3. Immobilized Human FGFR2 beta(IIIb) at 0.5μg/ml on the plate. Dose response curve for Anti-FGFR2 Ab., hFc Tag with the EC50 of 20.2ng/ml determined by ELISA.

Figure 4. Anti-FGFR2 Ab., hFc Tag can bind Human FGFR2 beta IIIb, His Tag with an affinity constant of 1.98nM as determined in a SPR assay (Biacore T200).

Human FGFR2 alpha (IIIb) Protein (FGR-HM1BD)

Figure 5. Serial dilutions of Anti-FGFR2 alpha (IIIb) Antibody were added into Human FGFR2 alpha (IIIb), His Tag : Biotinylated Human FGF10, No Tag binding reactioins.

Products

Background

Immune blockade therapy is becoming a new weapon in oncology due to the development of immune checkpoint proteins targeting in cancer, especially after the success of antibody drugs targeting programmed cell death-1 (PD-1), programmed cell death ligand-1 (PD-L1) and cytotoxic T-lymphocyte antigen-4 (CTLA-4). Although immune checkpoint blockade therapy often leads to a more durable response than chemo or targeted therapies in some cases, clinical data shows a limitation that in most cancers, the response rate is low (10-30%). Therefore, more associated studies focus on the mechanisms of non-responsiveness, as well as developing new targeting strategies using other immune checkpoint proteins.

Utilizing our featured SAMSTM platform, KACTUS has developed a series of immune checkpoint proteins with multiple pre-labels.

Background

FcR is the receptor of immunoglobulin (Ig), which causes corresponding cell-specific immune response by binding to the specific region of the Fc segment of Ig, acting as a bridge between humoral immunity and cellular immunity. Gene polymorphisms of FcR enable it to induce various intracellular biological effects, which are closely related to the occurrence and development of various immune-related diseases such as autoimmune diseases SLE and RA. Fc receptor proteins are one of the most important protein families in the immune system, mediating antibody-dependent ADCP, ADCC and CDC effects, and playing a key role in immune system activation and antibody function.

Currently the most widely used are mainly IgG receptors including FcRn and FcγR. FcyRs can be divided into three classes: FcyRI, FcyRII and FcyRIII. FcγRI has high affinity for IgG (10-9M) and is mainly expressed in monocyte-macrophages. FcγRII, FcγRIII and IgG have low affinity (10-6M) and are widely distributed. FcγRII is divided into 3 types: FcγRIIa, FcγRIIb and FcγRIIc according to the structure of the cytoplasmic region. It exists in B cells, macrophages, polymorphonuclear cells, platelets and monocytes, and FcγRIIb is the only inhibitory receptor. FcγRIII is present in granulocytes, NK cells, macrophages and activated monocytes. The combination of FcγR and antibody can activate immune cells and produce endocytosis, phagocytosis, ADCC and other functions.

Products

Background

CD47 is an immunoglobulin that is overexpressed on the surface of many types of cancer cells. CD47 forms a signaling complex with signal-regulatory protein α (SIRPα), enabling the escape of these cancer cells from macrophage-mediated phagocytosis. In recent years CD47 has been shown to be highly expressed by various types of solid tumors and to be associated with poor patient prognosis in various types of cancer.

A growing number of studies have since demonstrated that inhibiting the CD47-SIRPα signaling pathway promotes the adaptive immune response and enhances the phagocytosis of tumor cells by macrophages. Improved understanding in this field of research could lead to the development of novel and effective anti-tumor treatments that act through the inhibition of CD47 signaling in cancer cells.

Products

Transforming growth factor β (TGF-β) is a highly pleiotropic cytokine that plays an important role in wound healing, angiogenesis, immunoregulation and cancer. The cells of the immune system produce the TGF-β1 isoform, which exerts powerful anti-inflammatory functions and is a master regulator of the immune response.

Products