Abcore’s antibody discovery platform ends with delivery of positive, unique nanobody sequences. Often the next step in the process is to produce the nanobody for further testing. Depending on what further characterizations of the nanobody protein will be done, we can produce the protein in a few different purities. Currently all our protein expression is done in E. coli.
The first step is to grow a culture in volumes varying between 5 ml up to multiple liters. Then the E. coli undergoes osmotic shock, where we can isolate the nanobodies from the periplasmic space. This “shockate” is about 80-90% pure and is pure enough for many further experiments. However, because it is a mixture, it is hard to quantify the amount of actual nanobody present.
If the nanobody needs to be more pure or the concentrations need to be normalized for further experimentation, we can purify them more. In this case, Abcore purifies the nanobodies which have a His-tag, and we can purify them using a Ni-NTA (Nickel-NTA) chromatography. This results in a highly pure, known concentration of purified nanobody protein.