iCrea™ ADCC CD20 (+) Target Cell Line, Human Raji Cells

Description

iCrea™ ADCC Target CD20 (+) Assay Cell Line are designed to enable determination of the ADCC activity of existing or new anti-CD20 drug candidates.

iCrea™ ADCC Target CD20 (+) Assay Cell Line are based on a human B lymphocyte cell line, Raji ( CCL-86), and have been genetically engineered and optimized to express high and constant levels of the surface antigen CD20.

The cells are to be use as target cells for measuring the ADCC activity of anti-CD20 antibodies and are optimized to give high sensitivity and specificity when used together with iLite ADCC Effector (V) Assay Cell Line.

Features

Read-out of results: Quantitative
Detection system: Luminescence
Regulatory status: Research Use Only

Applications

The iCrea™ CD20 (+) Target Assay Cell Line can be used together with iCrea™ ADCC Effector (V) and iCrea™ CD20 (-) Target Assay Cell Line for the quantification ADCC activity of anti-CD20 antibodies.

Principle

iCrea™ADCC analysis preparation cells are modified cells, and antibody-dependent cell-mediated cytotoxicity (ADCC) can be checked by specific expression of firefly luciferase. When the target antibody binds to the antigen on the surface of the target cell, the target-binding antibody will be presented to the Fc receptor (FcγRIIIa) on the effector cell. When the Fc portion of the target binding antibody binds to the receptor, multiple cross-linking of the two cell types occurs. This initiates a signal cascade that triggers the expression of firefly luciferase (FL) in effector cells. In this application note, we describe the use of an effector cell line (iCrea™ ADCC Effector (V) Assay Cell Line) that overexpress FcγRIIIa and contains the FL reporter gene that mediates the signal from FcγRIIIa The main transcription factor transduced responds to the receptor, and the positive target cell line overexpressing the surface antigen HER2 (iCrea™HER2(+) target detection Cell Line). Under the control of a constitutive promoter, iCrea™ ADCC effector (V) analysis-Cell Line also contain the Renilla luciferase (RL) reporter gene, which allows the drug-induced FL activity relative to the constitutive expression of RL normalization. This makes the measurement result not affected by changes in the number of cells, serum matrix effects or target cell lysis effector cells. In addition, we described the use of a negative control as a target cell line depleted of negative cells (HER2(-) target detection Cell Line). After adding and incubating the luciferase substrate, the firefly luciferase signal can be measured in a luminometer. The firefly luciferase signal is directly proportional to the functional activity of trastuzumab in the sample (Figure 1).

Size

1 vial corresponding to one 96 well plate

Species Reactivity

Human

Cell Background

Raji cell line

Sample

Works with both adherent and suspension cells.

Growth Medium

Gibco (cat no 12648-010)

Cell Purity

>95%

Cell Viability

>90%

Sterility Testing

Creative Biolabs provides sterility testing in accordance with USP and EP regulations. All of our sterility testing is performed in an isolator or clean room environments. The cell line has been screened using the membrane filtration testing methods to confirm the absence of aerobic, anaerobic and fungi microorganisms.

Identity Testing

Identity testing is required for newly established cell lines. Isoenzyme analysis is used to confirm the identity of the species of a cell line. Alternative methods for identity testing include DNA fingerprinting, STR analysis and karyology.

Virological Safety Testing

A broad range of viruses is susceptible to affecting human cell lines. We can provide in vivo/vitro virus saftey assays by utilizing various animal systems. These viruses include: adventitious viruses, bovine viruses, human and simian viruses, porcine viruses, retrovirus and rodent viruses.

Genetic Stability Testing

We perform cell genetic stability studies under ICH guidelines. We can provide guidance on the appropriate testing program upon your requirements.

Mycoplasma Testing

Guaranteed mycoplasma-free

Shipping

Dry ice

Storage

Upon receipt confirm that adequate dry-ice is present, and the cells are
frozen. Immediately transfer to -80°C storage. Cells should be stored at
-80°C (do not store at any other temperature) and are stable as
supplied until the expiry date shown. Cells should be used within 30 min
of thawing.

Warnings

Avoid multiple freeze/thaw cycles

Research Use Only

Our recombinant cell are for research use only, not for diagnostic or therapeutic use.

Quality Control

The expression of hVISTA has been verified by flow cytometry.
The induction of antibody-dependent cellular cytotoxicity (ADCC) has been verified using anti-hVISTA-hIgG1 antibody and Jurkat-NFAT Lucia™ CD16 reporter cell line.
The stability of the 20 channels after thawing has been verified.
Raji-hVISTA cells are guaranteed to be free of mycoplasma.

Figure 1 Example of Trastuzumab calibration curve using the firefly luciferase substrate luciferase reagent.

Figure 2
A: Measurement of the specific Firefly (FL) signal. Cells were stimulated with increasing concentrations of Trastuzumab.
B: Measurement of the Renilla (RL) signal for normalization, from the identical assay as in A.

Figure 3 Dose response curve after normalization of the specific Firefly signal with the according Renilla signal.

iCrea™ ADCC CD20 (+) Target Cell Line, Human Raji Cells (CAT#: AD-P013-C) Datasheet

Overview

Specification

Images

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