Cell Based Assays

Antibody Dependent Cellular Cytoxicity (ADCC), Complement Dependent Cytoxicity (CDC), & Cell Mediated Cytoxicity (CMC)

 

We provide ADCC/CDC assay services to determine the activity of the antibody and/or complement against specified target cell line. the result would be a dose of dependent kinetic response of the therapeutic antibody. The assay service includes optimization of E/T ratios, measurement of ADCC/CDC activity at a series of antibody concentrations, at a certain E:T ratio. Correlation data with FACS analysis can be generated upon request.



Figure: 5000 Ramos cells/well were incubated with serially diluted Rituxan antibody for 15 minutes prior to the addition of purified NK cells stimulated overnight with IL-2. The ADCC reaction was further incubated for 2 hours (left graph) or 3.5 hours (right graph) at the specified E:T ratios. % Cytotoxicity was measured using the aCella TOX assay. 3.5 hour timepoint has been optimized with the 10:1 ratio (green). Graph with 20:1 ratio (red) is flat. (100% cytotoxicity)



5000 Daudi cells/wells were incubated with serially diluted Rituxan antibody for 15 minutes prior to the addition of purified NK cells maintained overnight with IL-2. The ADCC reaction was further incubated for 2 hours at the specified E:T ratios (25:1 black and 10:1 red). % cytotoxicity was measured using the aCella TOX assay.

The above graph shows an antibody titration of Rituximab from 0.000977 to 1ug/mL incubated with 5% rabbit complement and 5000 Daudi cells for 30 minutes at 37*C



Steps Involved in the ADCC/CDC Implementation:

1. Determination of optimal E:T ratios for the Target Cell Line
2. Procurement of the antibody (or antibodies) and Target Cell Line to be tested
3. A dose response curve would be generated using aCella TOX or FACS analysis or both (as requested)

Supplies & Information to be provided by the client:

1. Target Cell line(s) (adherent or suspension) (frozen vial(s)) and sufficient Antibody for the project
2.
Cell Culture Media and conditions for the growth of the target cell line Antibody concentrations (if known)
3. Number of PBMC donors to be tested
4. Information on Antigen expression level on the target cell line(s) (FACS analysis can be done at the request of the client to determine this)

Supplies to be provided by ImmunoChemistry Technologies:

1. All additional supplies would be procured by ICT and billed to the client at cost
2.
Final report would be provided after the project with a dose-response curve, showing Antibody dependence

Estimated Timeframe is approximately 4 weeks per cell line/antibody
For lead time and pricing, please call 1-800-824-8540 or email sales@immunochemistry.com

 

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