ADCC analysis of effector cell function identification and antibody drug development

A look at the battlefield of immune cells

Functional characterization of effector cells and antibody drug development of ADCC analysis

( Celigo Full Field Cell Scanning Analyzer)

1. ADCC function

ADCC acts as antibody-dependent cell-mediated cytotoxicity : refers to NK cells, macrophages, and neutrophils that express IgGFc receptors, by binding to virus-infected cells and The Fc segment of the IgG antibody on the surface of the target cell such as a tumor cell binds to the effect of killing these target cells.

2. The key point of ADCC action

1. Effective ratio (E:T): the ratio of effector cells to target cells is one of the key factors in the analysis of the role of ADCC.
2. Antibody concentration: the exploration of optimal antibody concentration is a key factor in the action of ADCC;
3. Effect time: the exploration of the best effect time point is also a key factor in the role of ADCC

3. Detection method:

The full-field cell scanning analyzer can monitor and quantify the killing battlefield of effector cells killing target cells at multiple time points.

4.Celigo monitoring experiment process example

1) Calcein AM Fluorescent Dyes: A cell staining reagent that fluorescently labels living cells.

Calcein AM can easily penetrate the living cell membrane. After entering the cytoplasm, the esterase will hydrolyze it into Calcein (calcium chlorophyll) and leave it in the cell, emitting strong green fluorescence. Because of its low cytotoxicity, it is the most suitable fluorescent probe for living cells.

1. At the time of analysis, the live target cells are labeled with Calcein, emitting strong green fluorescence; if the target cells die or lyse, Calcein is released and green fluorescence is no longer present. Therefore, the killing effect of effector cells on target cells can be analyzed by the change of Calcein AM fluorescence.

1.) ADCC analysis process:

5.Celigo imaging

1) 0h target cell full hole image green fluorescent image bright field + fluorescence superposition

2) 0h target cell count image (take partial image)

Green fluorescent cell image circled counting cell image

3) Target cell count images at different time points

0h 2h 4h 6h

6.ADCC killing formula and killing curve

CAM+_t0: Calcein AM positive number of 0h nCAM+_t: Calcein AM positive number of t hour

ADCC killing curve

7. Comparison of Calcein AM method and LDH release method based on Celigo image

Effect: (T E) to effect ADCC effector to target ratio different 8.

                  10:1 5:1 2.5:1

1.3:1 0.6:1 0.3:1       

9. Cell images of different effect time points

10. Cell images of different antibody concentrations

For details, please consult:
Daktronics for Biotechnology Co., Ltd. <br> Contact: Ms. Xiao Email:
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