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The preparation of medium for the cultivation in serum bottles demanded a different approach compared to the preparation of medium for the cultivation in benchtop bioreactors.
Preparation of medium was done on vertical autoclave (i.e., Robus Technologies).
The stock solution of 10 mM AgNPs was used for preparation of medium with AgNP concentration of 1, 3, 5, 7, and 9 nM.
High levels of nuclease activity may therefore only be a concern for a short period following preparation of medium from freshly thawed FBS.
Additionally, compared to traditional SSF, AF presents more advantages such as easier preparation of medium and more convenient product recovery [ 2].
Chemicals for preparation of medium and most other chemicals, including DAB, adenosine, glutamate, the glutamate receptor antagonist MPEP [2-methyl-6- phenylethynyl -pyridine], GABA and the ecto-ATPase inhibitor ARL67156 were purchased from Sigma.
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Basic skill experiment: This feature refers to the basic operation skills of a microbiology experiment, such as medical laboratory safety protection, preparation of culture medium, medium sterilisation, sterile working technique, operation of the microscope, bacteria dyeing, and bacteria separation.
At 20% confluence, replace D10 media with neuronal induction medium (see preparation of differentiation medium).
For preparation of conditioned medium (CM), protein concentration was determined using a Bradford assay; cells were cultured with serum-free medium (DMEM + 1% Pen/Strep) for 24 h and then concentrated using Vivaspin 20 Centrifugal Concentrating tubes (Sartorius, VS2021) at 3000×g up to 3 h until a desired concentration (10x) was reached.
An experimental design (Doehlert matrix) was drawn up to optimize the experimental conditions of the preparation of the medium.
Additionally, the NF retentate was reused for the preparation of culture medium, resulting in a change of the initial composition of broth.
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