I working with pancreatic cancer cell lines PT45P1 and would like to know whether there is a better test rather than wound healing assay to assess the cellular proliferation after and before treating the cells with chemotherapy. Also, I am interested to investigate how my drug effect the cellular apoptosis.
Thank you in advance
Wound healing assay is used to describe migration and proliferation simultaneously. Better way to addres that question is to perform MTT assay, BrdU incorporation assay, synthesis of ATP. If you want to check whether your drug induces apoptosis, you can do hoechst/propidium iodide staining, PARP cleavage on western blot, annexin/FITC analysis on flow cytometry or activation of effector caspases 3/7. Best luck!
Fawaz Al-Shaheri
you can use MTT assay to asses the proliferation and viability of cells
and use flow cytometry and annexin V test to asses apoptosis
also, bax/bcl-2 ratio, p53 and caspase-3 to evaluate apoptosis
You can combine MTT-test and acridine orange in same wells. Before the adding MTT in wells, add the AO (cells stained during 5-10 min, live - green, apoptotic - red/orange nucleus) and take imaging on microscope with UV-lamp, then add MTT for 4 h and then add 100 microL of DMSO in wells for dissolved cells and take spectrophotometry on 570 nm.
you can use flow cytometry and annexin V test to asses apoptosis and MTT ,LDH assay for proliferation and viability respectively.
Cell proliferation can be measured by MTT, XTT , SRB and trypan blue staining methods and For apoptosis determination, Use Annexin FITC/PI staining kit by flow cytometery or Fluorescent microscopy techniques. further you can confirm the apoptosis mode of cell death by western blotting analysis (cleaved PARP).
A BrdU incorporation assay, DiviTum, can be used in cell cultures, mouse and human sera/plasma. Dr Klaus Felix from Heidelberg have published results using DiviTum in pancreas PDAC: Article Preoperative Serum Thymidine Kinase Activity as Novel Monito...
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