ADCC Assays

Creative Biolabs is one of the forward-looking companies as well as undisputed market leader delivering impeccable services to customers all over the world. Currently, we offer this exclusive opportunity for our worldwide clients to adopt the most comprehensive ADCC assay service.

Introduction of ADCC

There are plenty of sophisticated mechanisms in the human immune system to identify and eliminate unwanted cells (e.g., viral infection and tumor). Amongst these, antibody-dependent cellular cytotoxicity, namely ADCC, plays a crucial role. After an antibody selectively coats the opsonized target cell, effector immune cells, such as natural killer (NK) cells, recognize and bind the Fc portion of the antibody, activating the effector cells to lyse the target eventually. ADCC represents one vital adaptive immune defense from infections or tumorigenesis in vivo. It is also considered as one of the most important mechanism-of-actions (MOAs) for therapeutic antibodies. Marketed therapeutic antibody products can deliver their clinical efficacy by induction of ADCC as the primary mechanism, or have such potential as their secondary or tertiary MOA. Therefore, to determine the exact ADCC liability and subsequent measurement, if present, constitutes an imperative step in the development and functional characterization of antibody therapies.

Fig.1 Significant differences among the ADCC examinations.^{1, 2}

Equipped with world-class technologies and seasoned team of experts, Creative Biolabs has developed, optimized, and validated an integrated ADCC evaluation platform. We provide multiple assay formats, from which you can select the best fit for your specific project.

Target Cells

We can deal with frozen cells from customers upon request. In case you haven't got an ideal target cell line, you can rely on our extensive expertise to find the most appropriate one. There are over a hundred well-qualified tumor cell lines available at Creative Biolabs. Our scientists also have vast experience in designing a variety of disease models for in vivo pharmacology research.

Effector Cell

NK cell is the major effector immune cell involved in ADCC, which can interact with Fc portions of IgGs via Fc receptor, mostly FcγRIIIa (CD16a) on its surface. Notably, human FcγRIIIa displays a dimorphism in the position of residue 158, resulting in a higher affinity variant (V variant) and a lower affinity variant (F variant). Given the essential role of effector cells in ADCC, Creative Biolabs provides multiple choices for best assay performance.

• Primary ADCC Assays - Primary peripheral blood mononuclear cells (PBMCs) or isolated NK cells are classical effector cells that are widely used in ADCC assays. Creative Biolabs can harvest fresh PBMCs or purified NK cells from healthy or diseased donors, with different genotypes (FF/VV/FV), by standard procedure. For primary NK cells, we apply strict quality control by screening CD16/CD56 marker expression using flow cytometry. A battery of samples from different sources can help to minimize donor-specific variability.
• ADCC Reporter Assay - ADCC assays with primary effector cells can be time-consuming and expensive. Thus, we provide alternative approaches, namely ADCC reporter bioassay, in which engineered effector cells stably expressing FcγRIIIa and luciferase are used.
• Mouse ADCC Reporter Bioassay
  Creative Biolabs provides a mouse ADCC reporter bioassay that employs the mouse FcRIV-expressing effector cell and luciferase to evaluate the proposed antibody. We have the capacity to effectively accomplish projects for customers all around the world while assuring their satisfaction.

Assay Endpoint

• Target cell lysis: LDH (lactate dehydrogenase) assay/ATP release/FACS screening
• Effector cell activation: luminescent NFAT reporter
• Real-time monitoring: instead of single-endpoint measurement, this platform can dynamically analyze target cell proliferation and death in the presence of effector cells and testing antibodies, thus allowing simple and sensitive detection of subtle ADCC activities.

Published Data

Data: This investigation examined the immunological potential of antibody-dependent cellular cytotoxicity (ADCC) triggered by two hemagglutinin (HA) head epitopes (E1, E2) in pandemic H1N1 influenza as a basis for broad-spectrum vaccine development. Experimental data demonstrate that E1 immunization provoked robust ADCC responses in vitro and in murine models, accompanied by moderate viral titer reduction. Paradoxically, E1-vaccinated hosts exhibited exacerbated pulmonary pathology and elevated mortality post-infection, a phenomenon potentially linked to dysregulated pro-inflammatory cascades.

Method: Flow cytometry-based assay

Fig.2 Sera of E1-vaccinated mice improve responses in ADCC.^2,4

Data 2: This study created an ADCC assay using target cells expressing MERS-CoV spike protein and effector cells engineered to express FcγRIIIA and a reporter gene. The assay measures reporter gene activity (luciferase) after anti-MERS antibody-mediated interaction between target and effector cells. The method was optimized following established guidelines. The assay effectively measured anti-MERS antibody ADCC activity. Notably, antibodies lacking fucose (afucosylated) demonstrated greater ADCC activity than those with fucose, suggesting that this modification can improve antibody-mediated viral clearance.

Method: ADCC reporter assay

Fig.3 Evaluation of antibody biological activity as assessed through ADCC reporter assay versus cytotoxicity assay.^3,4

Based on unparalleled technologies and abundant resources, this robust and reliable ADCC test system of Creative Biolabs not only guarantees the most accurate and reproducible outcomes but also accommodates any custom experimental designs. Our highly specific assays can offer absolute confidence in evaluating clinical effects and risk potentials concerning ADCC activity.

Besides, Creative Biolabs also offers complement-dependent cellular cytotoxicity (CDC), antibody-dependent cellular phagocytosis (ADCP) for investigating the antibodies’ mechanism of action (MOA), as well as to ensure efficacy and safety.

For more detailed information, please feel free to contact us or directly sent us an inquiry.

References

1. Lewis, George K., et al. "Knowns and unknowns of assaying antibody-dependent cell-mediated cytotoxicity against HIV-1." Frontiers in Immunology 10 (2019): 1025. Distributed under Open Access License CC BY 4.0, without modification.
2. Ye, Zi-Wei, et al. "Antibody-dependent cell-mediated cytotoxicity epitopes on the hemagglutinin head region of pandemic H1N1 influenza virus play detrimental roles in H1N1-infected mice." Frontiers in Immunology 8 (2017): 317.
3. Cao, Junxia, et al. "Development of an antibody-dependent cellular cytotoxicity reporter assay for measuring anti-Middle East Respiratory Syndrome antibody bioactivity." Scientific Reports 10.1 (2020): 16615.
4. Distributed under Open Access License CC BY 4.0, without modification.

For Research Use Only.