Product Number CM1xxxx
CellMosaic’s product number (catalog number or SKU) CM1xxxx is generally assigned to any small molecules and their bioconjugation kits such as drug molecules, drug intermediates, crosslinking reagents (oxLink™ and sxLink™), and antibody drug conjugation kits and other drug conjugation kits.
CM11409: Antibody MMAE Conjugation Kit (with VC-PAB linkage)
Can CM11409 be used to couple a scFv (single chain variable fragment)?
CM11409 uses a reducing reagent to partially break the disulfide bonds linking two heavy chains. CM11409 can be used depending on how the fragments of the scFv are linked but, unless you know the structure of the disulfide, it is not recommended. Instead, we recommend using CM11413 to label the scFv. This kit uses the surface amine of the protein to generate the thiols needed for linking to Mc-VC-PAB-MMAE. MMAE can be released from the ADC the same way as the ADC prepared using CM11409.
For the DAR estimation by UV, what are the parameters of 1.615, 0.1425, and 9 in your protocol? Could I use this formula for any antibody?
These parameters are derived from the extinction coefficient of the antibody and drug. The formula is only applicable to a standard IgG assuming a MW of 150 KDa and extinction coefficient of 210,000. If your antibody is not a full length, you can use the calculation sheet from CM11413 (https://www.cellmosaic.com/protein-mmae-conjugation-kit/)
Do you have any suggestions to minimize aggregation during the conjugation process. Can we add a little Tween 20 or Tween 80 to the reducing buffer, labeling buffer, or storage/elution buffer (PBS) to prevent aggregation?
The aggregation is mainly caused by the hydrophobicity of the drugs, possibly due to stacking. To decrease the aggregation, you will have to use a water-soluble drug or a drug with a less hydrophobic linker, such as our super hydrophilic AqueaTetherÒ (AqTÒ) linker. For more information on this linker, please see https://www.cellmosaic.com/aqueatether-aqt-technologies/.
You can also reduce aggregation by decreasing the number of drugs loaded per antibody. Adding detergent may not help in this case because it is not like a membrane protein, and micelle formation is promoted. However, you are welcome to try adding detergent to the storage buffer after purification. We do not recommend adding detergent to the reducing buffer and labeling buffer because this affect the reaction.
We do have ADC stabilizing PBS buffer that you can use for long-term storage of ADC: https://www.cellmosaic.com/adc-stabilizing-buffer/ The buffer can keep the aggregation low during storage.
CM11416: F(ab')2 MMAE Conjugation Kit (with VC-PAB linkage)
What MW of conjugate can I expect to see under non-reducing SDS-PAGE? Will the final products still be intact after the reduction? If the molecule falls apart during the reduction, will a reoxidation step after conjugation help put the V/L chains together?
You should expect to see ~110 KDa product after conjugation under non-reducing conditions. The reducing condition is mild and will partially reduce the F(ab’)2, so F(ab’)2 should remain intact. There is no reoxidation step because the reduced thiol will react with the drug and form the conjugate. Depending on how your F(ab’)2 is prepared and how easy the heavy chain can be reduced, you may see a small percentage of the F(ab’)2 falling off (note: we have not seen it before). If that is the case, you may have to use a diffident chemistry (kit) to prepare the conjugate.
CM11431: Antibody Deruxtecan Conjugation Kit
We would like to use this kit for scFv-Fc conjugation (MW = 110 kDa, 2 disulfide bonds). Do you think you could customize the protocol for me?
Yes, we can customize the protocol to accommodate the need. You can select “Need Customization (1-3 mg None IgG Type Antibody)” (AS1004) under the Antibody PerKit™ Customization when you place the order online.
CM11425: Antibody MMAF Conjugation Kit (with VC-PAB linker)
I just received the MMAF kit, the gel pack inside the packaging and the MMAF was no longer cold. Do you think this could change the quality of the MMAF?
Box 1 contains Mc-VC-PAB-MMAF and Reagent A which are relatively stable at room temperature for a few weeks. They are usually shipped at room temperature by other vendors. We use a cold pack to ensure the temperature does not rise during shipment. The quality of the MMAF should be fine.