August 3, 2020
by Thomas Porter
Lab activities are back to a new normal and we have restarted cell culture for labeling using DNA cages. The first step in this process is optimizing DNA conjugation to antibodies. The process being used is depicted below.
Every step of the process was optimized starting with replacement of the DBCO-NHS used and the reduction the amount of DBCO used by 10-fold. The original amount of DBCO used likely lead to cross-linking between the antibodies. Once the amount of DBCO was decided, the amount of excess DNA used was also reduced. After the optimization process the concentration of antibodies recovered after conjugation is 2-3 times higher and the conjugation efficiency is now approximately 10% as compared to the previous ~2.6%. The conjugation efficiency is calculated by dividing the molarity of the antibodies by the molarity of the fluorescent DNA which binds to the DNA on the antibodies which may not be the most accurate calculation method.