Laleh

Dear Friends We are going to isolate human mesenchymal stem cells from fat tissue. Different studies have used different media for isolation and expansion of these cells. Which medium is more suitable for these cells? DMEM high glucose, DMEM low glucose, DMEM/F12 or alpha MEM all supplemented with 10-15 percent FBS. Any suggestions will be appreciated. Regards

Dear CharonY Thank you very much for your comments. I will certainly repeat the experiment and increase the replicates, while checking the quality of RNA and melt curve more carefully. Thanks a lot

Thank you

Dear CharonY Thank you very much for your comments. We use HighRox SYBR green mix on ABI stepone plus I have included Cts here. Please take a look: Cell on scaffold (test): Ct1 for gene of interest: 34, Ct2 for gene of interest: 33. Ct1 for GAPDH: 31, Ct2 for GAPDH: 31 Control cells (without scaffold): Ct1 for gene of interest: 32, Ct2 for gene of interest: 32. Ct1 for GAPDH: 21, Ct2 for GAPDH: 22 I do not have access to the melt curves right now In your opinion what causes the huge difference in Cts of GAPDH between samples? Thanks a lot

Hi everyone I have performed an relative quantification on two different samples one cultured cells, one my cells on a tissue engineering scaffold. I have had two replicates in each real time PCR assay, and conducted the experiment two times. while in my control group which only consists of cells the Ct for house keeping gene-GAPDH-is relatively low-around 21- the Ct for GAPDH is around 31 in my test group-cells on scaffold. I have attached the Ct results. I was wondering if this result can be valid or scaffold somehow affects GAPDH expression? please note that the Ct for my gene of interest does not have much difference between test and control. Cts.docx