What one change to your room, system, or setup made the biggest improvement?

Hard to assign true impact differentials, since all these upgrades work cumulatively.

It might be hard to be exact, but why it would be hard to estimate the difference made by larger changes is unclear. The difference between a room with flabby bass or echoes is a huge change, and the fact that, overall, a series of changes are cumulative does not prevent that observation.

@hilde45 

From my experience, because of 2 (related) factors:

• ordinality & 2nd order effects: some improvements only shine once others are in place. E.g. the impact from the Townshend podia in my room increased notably after I had more room treatment. 
• discrete, step function rather than continuous increases. Once a combination of upgrades (power, cables, ...) jointly lowered the noise floor below a certain threshold, suddenly instrument separation drastically improved. 

Perhaps more experienced audiophiles can factor these interactions into their upgrade path and disambiguate their individual vs cumulative/interactive effect, but for me there were clear moments where the effect of 'the whole' was greater than that of the sum of its parts. I am very much still in the empirical rather than the rationalist phase of system optimization.  

Speakers equipped with top quality drivers. 

Pulling speakers way out into the room. The entire space between speakers and rear wall became the soundstage. 

The first was adding room treatments, both absorption and diffusion and then it was going from 2 to 4 subs.