Travelling guitar.

[impala59]

If it's a tele or Strat style fixed bridge, you want to measure to the high E string saddle. The saddle should be close to it's furthest amount of travel towards the nut, but not fully maxed incase you get any fine measurements wrong and need to compensate down the track.

The idea is the low E always sits back the most in common intonation and the other strings incrementally in between. This pic should explain the general shape of the saddles

Does that always apply with 25.5" scale?
Is it always best to set the bridge position using the high E as datum?
What if using a shorter scale?
Sorry if asking basic questions but am only recently getting involved in building. Is there a formula to follow? I ask as I have built some 34" scale basses and maybe just got lucky with positioning and subsequent intonation

[Bakersdozen]

Does that always apply with 25.5" scale?
Is it always best to set the bridge position using the high E as datum?
What if using a shorter scale?

Yes, as the other strings traditionally intonate further back.

You will see a lot of tune-o-matic style bridges are angled, with the low E the furthest back from the nut. Also some adjustable saddle bridges (think Strat and tele style) come with the adjustment screws in staggered lengths, with the shortest screws toward the bass end.

Same deal with any size scale lengths. Just remember to have the high E saddle close to the front end of travel, but not fully maxed out just in case. If you have a decent search, there are heaps of posts in here going into further detail on the matter. I remember Mr @Simon Barden has written a few great in-depth posts about this.