I am culturing embryonic neural progenitors cell from mice from E11.5 time pregnant mice. And differentiating them into neurons by removal of bFGF but after doing qPCR of cells without bFGF and Neural Precursor cells in media containing bfgf the change in sox2 and nestin level is not prominant but change in tuj 1 is huge what could be possible cause?
Supposed your controls are OK, and also that qPCR may reflect somehow the protein level in those cells, I could think of already too much differentiated cells, wich might be consistent with high levels of tuj1 and only low levels of nestin, as you observed. This may also correlate with low sox2. Perhaps you need a different (smaller) time window?
I also got the same problem with human Neural Stem Cell (hNSC).
I tried to differentiate hNSC follow the protocol provide by Thermofisher (https://www.thermofisher.com/jp/en/home/references/protocols/neurobiology/neurobiology-protocols/differentiating-neural-stem-cells-into-neurons-and-glial-cells.html).
However, after 4, 6, and 8 weeks of differentiation, immunoflourescence staining with Nestin and Tuj1, I found that Tuj1 is up-regulated but the maybe no change in the level of Nestin, but I found that there are cells negative for Nestin. Later, I saw that in the protocol, they only observed for neuronal marker after differentiation but not Nestin/Sox2.
Checking a paper from Zhang and Jiao, I think that maybe it is possible for differentiated cells to express Nestin beside Tuj1 or DCX (please check the paper here https://www.hindawi.com/journals/bmri/2015/727542/#B60). However, I do not know whether my thought is correct or not.
Dear Vuong are you checking Level of Rest also and yes , did you Noticed any change in its level.
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