Anti-Human VEGF (Bevacizumab) – Fc Muted™ Biotin

Anti-Human VEGF (Bevacizumab) – Fc Muted™ Biotin

Product No.: LT406

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Product No.LT406
Clone
A4.6.1
Target
VEGF
Product Type
Biosimilar Recombinant Human Monoclonal Antibody
Alternate Names
Vascular Endothelial Growth Factor; VEGF-A; VEGFA; Vascular Permeability Factor; VPF
Isotype
Human IgG1κ
Applications
ELISA
,
FC

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Antibody Details

Product Details

Reactive Species
Human
Host Species
Human
Expression Host
HEK-293 Cells
FC Effector Activity
Muted
Immunogen
Recombinant human VEGF.
Product Concentration
0.5 mg/ml
Formulation
This Biotinylated antibody is formulated in 0.01 M phosphate buffered saline (150 mM NaCl) PBS pH 7.4, 1% BSA and 0.09% sodium azide as a preservative.
Storage and Handling
This biotinylated antibody is stable when stored at 2-8°C. Do not freeze.
Regulatory Status
Research Use Only (RUO). Non-Therapeutic.
Country of Origin
USA
Shipping
Next Day 2-8°C
Applications and Recommended Usage?
Quality Tested by Leinco
FC The suggested concentration for Adalimumab biosimilar antibody for staining cells in flow cytometry is ≤ 1.0 μg per 106 cells in a volume of 100 μl. Titration of the reagent is recommended for optimal performance for each application.
ELISA
Additional Reported Applications For Relevant Conjugates ?
B
N
IP
WB
Each investigator should determine their own optimal working dilution for specific applications. See directions on lot specific datasheets, as information may periodically change.

Description

Description

Specificity
This non-therapeutic biosimilar antibody uses the same variable region sequence as the therapeutic antibody Bevacizumab. Bevacizumab recognizes both native and reduced human VEGF (isoform 165). This product is for research use only.
Background
Bevacizumab is a monoclonal antibody that specifically recognizes vascular endothelial growth factor (VEGF). VEGF is a growth factor that participates in angiogenesis, vasculogenesis, and endothelial cell growth. It facilitates endothelial cell proliferation, cell migration, and the permeabilization of blood vessels. In addition, VEGF inhibits apoptosis. Bevacizumab neutralizes the biological activity of VEGF by preventing the interaction of VEGF with its receptors on the surface of endothelial cells, resulting in the regression of tumor vascularization, normalization of remaining tumor vasculature, and inhibition of the formation of new tumor vasculature, thus inhibiting tumor growth.1 Anti-Human VEGF (Bevacizumab) utilizes the same variable regions from the therapeutic antibody Bevacizumab making it ideal for research projects.
Antigen Distribution
VEGF is widely expressed in the thyroid, prostate, and various other tissues.
PubMed
NCBI Gene Bank ID
Research Area
Biosimilars

Leinco Antibody Advisor

Powered by AI: AI is experimental and still learning how to provide the best assistance. It may occasionally generate incorrect or incomplete responses. Please do not rely solely on its recommendations when making purchasing decisions or designing experiments.

Research-grade Bevacizumab biosimilars are used as calibration standards or reference controls in pharmacokinetic (PK) bridging ELISA assays to provide accurate quantification of drug concentrations in serum samples, enabling consistent and comparable results across assays and laboratories.

  • Calibration Standards: In a PK ELISA, a standard curve is generated using known concentrations of Bevacizumab or its biosimilar form, diluted in the same matrix as the test samples (e.g., serum). These standards fully cover the expected range of drug concentrations, which may span from values such as 39.06 ng/mL to 2500 ng/mL depending on the kit or protocol. The absorbance (signal) produced by each standard concentration creates a reference curve that allows unknown sample concentrations to be interpolated.

  • Reference Controls: Biosimilar Bevacizumab preparations are also used as reference controls, especially for assay validation or bridging studies when switching assays, comparing results across laboratories, or when harmonizing studies globally. Controls of known concentration are run alongside samples to confirm the ELISA’s accuracy and precision.

  • Bridging Studies and Harmonization: The availability of internationally recognized standards, like the 1st WHO International Standard (IS) for Bevacizumab (coded 18/210), supports the calibration of secondary or in-house standards used in local laboratory assays. Using a well-characterized biosimilar as reference minimizes inter-laboratory variability and enables harmonized quantification and biological activity assessment across different Bevacizumab products and platforms.

  • Biosimilars as Equivalents to Originator in ELISA: Research-grade biosimilars are formulated to be pharmacologically and structurally equivalent to the reference Bevacizumab (e.g., Avastin). This equivalence allows them to serve interchangeably with the originator in assay calibration and controls, provided their bioactivity and binding characteristics are confirmed.

  • Validation Guidance: PK ELISAs using such standards are developed and validated in accordance with regulatory guidance (e.g., ICH M10, FDA), ensuring that the assay’s linearity, accuracy, and precision are established for all relevant matrices and sample handling conditions.

Key Points:

  • Calibration standard curves are constructed using research-grade biosimilars diluted in serum or sample buffer.
  • Reference controls (biosimilar at known concentrations) are run to validate assay performance.
  • WHO International Standards help harmonize results, reduce assay/batch variability, and standardize potency assessments for biosimilar comparability.

This approach ensures patient samples’ Bevacizumab (originator or biosimilar) concentrations can be reliably quantified for PK analysis and regulatory submissions.

Direct flow cytometry protocols for PE or APC-conjugated Bevacizumab biosimilars to assess VEGF target expression or binding are not described in detail in the search results, but standardized approaches can be reliably extrapolated from related monoclonal antibody biosimilar validation workflows. The typical protocol structure for such assays is as follows:

Essential protocol steps:

  • Cell Preparation
    Culture and harvest the cell line expressing the VEGF target. Wash cells in cold PBS or similar buffer to minimize nonspecific binding.

  • Antibody Incubation
    Incubate the cells with a titration series of the PE or APC-conjugated Bevacizumab biosimilar in cold buffer (usually 4°C, to prevent internalization), for 30-60 minutes. Titration ranges often span from ~20 μg/mL down to 0.04 μg/mL, depending on expected affinity. No secondary antibody is required because the biosimilar is directly labeled.

  • Wash Steps
    Wash cells thoroughly (often 1-3 times with cold buffer) to remove unbound antibody conjugate.

  • Flow Cytometry Acquisition
    Analyze cells on a flow cytometer equipped for the relevant fluorochrome (e.g., PE or APC detection).

  • Data Analysis
    Quantify mean fluorescence intensity (MFI) as a surrogate for VEGF expression level or binding capacity. Plot dose-response curves (e.g., 4-parameter logistic, 4-PL) to compare saturation, affinity, or capacity between test biosimilars and reference molecules. Use statistical methods (e.g., ANOVA, parallel line analysis) to assess binding similarity and potency between biosimilar and reference standards.

Protocol schematic adapted for Bevacizumab biosimilar validation (based on ):

1. Harvest 1 × 10^5 cells and resuspend in 50 μL cold PBS.2. Add conjugated biosimilar (range: 20–0.04 μg/mL).3. Incubate for 30 min at 4°C.4. Wash cells once or twice in cold PBS.5. Resuspend in 200 μL PBS and acquire on flow cytometer.6. Analyze MFI, plot dose-response, and assess statistical equivalence.

Key considerations for validation:

  • Use an excess of cells to ensure robust quantification.
  • Include unstained, isotype, and reference standard controls to benchmark specificity and sensitivity.
  • The approach is directly analogous to protocols for trastuzumab biosimilar validation, as both rely on direct antibody-antigen interactions measured by flow cytometric fluorescence.

Additional points:

  • If unconjugated biosimilars are used, an additional incubation step with a labeled anti-human IgG secondary antibody is required.
  • While not shown for Bevacizumab in the search results, this protocol structure is recognized as industry standard for flow cytometric biosimilar binding validation.

Limitations:

  • Specific published protocols for PE/APC-conjugated Bevacizumab biosimilars on VEGF-expressing cells were not identified in the provided search results, so recommendations are based on robust inference from closely related mAb biosimilar flow cytometry protocols.

Summary:
A standard protocol to validate VEGF expression or binding of a PE/APC-labeled Bevacizumab biosimilar by flow cytometry involves direct incubation of the labeled antibody with target cells, followed by acquisition and quantitative analysis of fluorescence intensity—mirroring procedures for other mAb biosimilars such as trastuzumab.

Biopharma companies verify the structural and functional similarity of a proposed biosimilar to the originator (reference) drug using a comprehensive suite of analytical assays designed to compare protein structure, physicochemical properties, biological activity, and impurity profiles. The Leinco biosimilar typically serves as a reference standard, assay control, or reagent in these analytical studies, enabling side-by-side comparisons.

Key Analytical Assays for Structural Similarity:

  • Primary structure: Mass spectrometry (MS), peptide mapping (to confirm amino acid sequence).
  • Higher-order structure: Circular dichroism (CD), nuclear magnetic resonance (NMR), X-ray crystallography, and differential scanning calorimetry (DSC), which analyze protein folding and conformational integrity.
  • Post-translational modifications: Glycosylation profiling by MS, capillary electrophoresis (CE), or chromatography to detect pattern similarities (critical for biologics such as monoclonal antibodies).
  • Physicochemical properties: Isoelectric focusing, HPLC (size-exclusion, reverse phase), SDS-PAGE for charge and size variants and to detect aggregates or fragments.

Key Analytical Assays for Functional Similarity:

  • Biological assays: Cell-based potency assays measuring intended biological activity (e.g., antibody-dependent cell-mediated cytotoxicity [ADCC], complement-dependent cytotoxicity [CDC]).
  • Binding assays: ELISA, surface plasmon resonance (SPR), or Biacore for antigen, receptor, or Fc receptor binding.
  • Enzyme kinetics or activity assays when relevant to the mode of action.

Impurity and Purity Analysis:

  • Impurity profiles: Analytical assessment of process-related and product-related impurities, using orthogonal chromatographic and electrophoretic methods. This confirms that impurity levels are comparable and do not introduce safety concerns.

Assay Controls and Reference Standards:

  • The Leinco biosimilar is frequently used as a side-by-side comparator or as an assay reagent in analytical and functional tests to directly compare its attributes with those of the originator drug and control standards. It may serve as:
    • Reference material for spiking, calibration curves, or assay validation.
    • A comparator in head-to-head structural, binding, or potency studies.

Purpose and Importance:

  • These assays ensure that critical quality attributes (CQAs)—the structural and functional characteristics most likely to affect clinical outcomes—are highly similar between the biosimilar and the reference product, which is a central requirement for regulatory approval.
  • The use of multiple, orthogonal (complementary) techniques provides robust evidence and can sensitively detect even minor differences, which are then evaluated for potential clinical relevance.

Altogether, the analytical comparability exercise integrates these platforms to fingerprint the biosimilar relative to the originator, confirming that any differences detected are not clinically meaningful. The Leinco biosimilar, as a high-quality comparator or assay reagent, is critical for ensuring assay reliability and meaningful comparison outcomes.

References & Citations

1. Pazdur, R. et al. (2018) Clin Cancer Res. 24(18):4365-70.
Indirect Elisa Protocol
Flow Cytometry

Certificate of Analysis

Formats Available

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Disclaimer AlertProducts are for research use only. Not for use in diagnostic or therapeutic procedures.