This is hopefully a really basic error I'm making, but I am very new to this.
I'm using the TaqMan Gene Expression Assay to measure thrombomodulin expression in endothelial cell lysates. I'm using 18s as the endogenous control. My results are all over the place. Even untreated samples return hugely different dCt values (4 cycles apart). The strange thing is that the curves for the thrombomodulin probe are always pretty much superimposed. All the variation comes from the 18s curves which are widely spread (4 cycles apart!)
I can accept that the 18s curves will separate a little due to pipetting error, but it seems a mighty coincidence that every time the thrombomodulin curves are closely packed.
I hope this makes sense to someone!