thanks a lot Piyush,

Normal medium ist with or without Penicillin-Streptomycin ?

any medium used for stimulation should contain the following: Steptomycin/Penicillin,

L-Glutamin, FCS. Stimulation can be varied between 24-72 hours according to the cell type and allergen used (OVA, HDM extract, Birth Extract) you can see all the papers published by group of Prof. Harald Renz (My Supervisior) which will help you alot i hope.

It depends on the output you want to analyze. For example to access the effect of your stimulus on the activation of intracellular signaling cascades (p-38, ERK, JNK etc…) you will obtain better results by starving the cells for several hours. This is because serum contains grow factors that activate some of these pathways leading to elevated phosphorilation levels even in untreated cells. Therefore you can starve the cells for several hours and then add your stimulus in complete media.

You refer that your stimulus is an allergen. Cutaneous or respiratory? In those cases it could be beneficial to have serum in the media when you add your allergen. Sensitizers react with serum proteins to form haptens that interact therefore with immune cells such as dendritic cells etc.….

For ILs expression, you should growth cell line in the media complete (+FBS). And for improve ILs concentration, the media should content LPS aslo. Good luck brother

Hi Dung. Sorry, but I don´t understand why do you want stimulate a cell line with an allergen. Generally, allergen stimulation is used to see a specific effect, example specific proliferation or specific cytokine production. What type of cells do you use? Regarding your question, to measure cytokines is necessary that the cells are in culture by several hours or days, so cells need all nutrients (serum, L-glutamine, pyruvate in several cases) and is appropiate use also an antibiotic to avoid contaminations