Hi Ranu, I suggest you read the paper: Analyzing real-time PCR data by the comparativeCT method (Schmittgen and Livak, 2008). I hope it helps you, there they explain very well the different methods to calculate the relative expression and the fold-change, with clear examples.

Hi Ranu

There is some difficult to determine in the immediate post-operative

You can check this reference

"Pre-Operative Assessment and Post-Operative Care in Elective Shoulder Surgery"

Article Pre-Operative Assessment and Post-Operative Care in Elective...

"Pre-Op" is your reference sample.

The term "relative expression" is not clearly defined and should not be used. If it is used, it should be made clear relative to what the measure is given. The fold-change (A/B) is a relative expression (the expression in A relative to the expression in B).

Many authors term 2^(dCt) as "relative expression" (with the intendet meaning: the expression of gene X relative to the expression of a gene Y in a sample), what is not correct. Since the two genes are measured with different assays (primers, threshold), 2^(dCt) is a product of the relative expression and some unknown (assay-specific) factor. It's better formany reasons to avoid 2^(dCt) and simply present dCt values directly.

If the data are paired (as it could be here, when a dCt is determined pre-OP and post-OP from the same individual), one would calculate an individual ddCt per individual. Although 2^(ddCt) now is in fact a fold-change (because the assay-specific unknown constant now cancels out), I recomment still just presenting the ddCt values, as they are (i) closer related to the biological relevance of the effects, (ii) they keep the symmetry of the problem (an induction in A/B should correspond to a similar repression in B/A; in the ddCt scale, this is reflected just by the sign: if A/B is +3, the B/A is -3: same size but opposite direction), and (iii) statistics are simpler, because ddCt values are approximately normal distributed, but fold-changes are severely right-skewed. This goes all confused and wrong in Livaks paper (referenced by Mitzuko Dautt-Castro above). Additionally, Livak misses to recognize that Ct-values are proportional to minus log(Conc), leading to an unfortunate order of subtractions of Ct-values, the difficulty to interpret dCt values and the need to correct the sign at the end (they need to calculate FC= 2^(-ddCt); mind the minus sign in the exponent).

Hi Ranu

You can method in this paper

A combination of circulating miRNAs for the early detection of ovarian cancer

"Article A combination of circulating miRNAs for the early detection ...

Thanks for answer my question Mr. Laith, Jochen, and Ms. mitzuko..