A friend posed a question to me last night. I have my own opinion and I
gave it to him. I would be interested in hearing what other scientists
might think on the matter:
I will change the details a little just to protect the innocent.
The experiment being conducted measure the effect of a particular drug on
the levels of certain liver enzymes. The levels of the enzymes will be
quantitated by western blotting.
In the experiment, 8 mice are treated with the drug for the prescribed
time and dosage and 8 mice are treated with vehicle. Their livers are
pooled according to treatment group (drug or control) and homogenized.
SDS-PAGE is done and the
blots are performed.
Now, if the scientist runs a 7 lane gel, where:
lane 1 = MW markers
lane 2, 4, 6 = control homogenate
lane 3, 5, 7 = drug treated homogenate
and then blots the entire piece of nitrocellulose for the level of target
enzyme, is that 3 replicate measurements (comparing lane 2 vs. 3; 4 vs. 5;
6 vs. 7) ?? Or is that an n of 3?
or second scenario:
if the scientist runs a 3 lane gel, where:
lane 1 = MW markers
lane 2 = control homogenate
lane 3 = drug treated homogenate
and blots for the enzyme. And then runs a second identical gel next week and
blots that; and then runs a third identical gel the following week and
blots that; is that 3 replicate measurements (remember, the very same
homogenates
were used in all the blots), or is that an n of 3??
The answer to me seems pretty obvious in either scenario, but I'd like to
hear other opinions too.
John
hines at pharm.med.upenn.edu
