Hi
Dear all
I used aggregated amyloid beta 1-42 (10 micromolar) to induce Alzheimer's condition in PC12 cell line, but after 24 h incubation, MTT assay was done, I obtained 85% cell viability, I have a question and I would like to guide me?
is it possible to achieve the alterations such as hyperphophorylated proteins using molecular analysis like western blotting or RT-PCR while we did not observe cell death (50%) using MTT assay?
I'm looking forward to getting your answer,
Thanks a lot,
Bets wishes
Marzieh
qRT-PCT ist not useful for phosphorylation analyses, this should be done via Western blot analyses.
Thanks a lot for your kind answer and waiting for useful giudance
I want to know why 10 micromolar of aggregated amyloid beta did not show cell death (50%), and if I do not consider the results of MTT assay and several proteins are investigated using western blotting, I may be obtain valuable and nice data?
Thanks a lot
Marzieh
In general - and if results of otheres (50% reduced viability") are reliable - WB will not be an adequate compensation for mising viability data. However, you could perform alternative viablility assays, such as Calcein AM assay, or poptosis testing (AVPI staining), or analyses og Bcl-2 vs BAX expression etc. to determine viability, apoptosis and/or surcival of cells. This depends on the functions you want to determine, such as cell proliferation (MTT or CFSE staining), cell viability (calcein AM assay), apoptosis/survival (expression Bcl2/BAX or better AVPI assay (FACS method)).
Thanks a lot for your useful and complete answer,
I want to assess several parameters resulting from the induction of Alzhemer's, so it is possible to observe your mentioned alterations although I did not observe cell death using MTT?
I'm looking forward to getting your answer
Best wishes
Thanks a lot
Marzieh
if the stimulus works this (at least in part) should be visible.
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