There is little point in setting your microsteps as high as 3200.
My recommendation would be that if you are Metric then use 1/10th microstepping (2000) or if you are Imperial use 1/8th microstepping (1600).
If you are using the Parallel Port then select that (do not use Max NC Mode), also set Kernel Speed to 25000Hz. If you are not using the parallel port but using a motion controller then which type ?
For distance calibration, initially, make sure you are setting-up Mach3 in the Units (Inch/mm) you intend to use (Config. / Set Native Units).
In Members documents there is a calculator for finding Steps per Unit and also a math's example.
http://www.machsupport.com/forum/index.php/topic,16315.0.htmlhttp://www.machsupport.com/forum/index.php/topic,12512.0.htmlThere is also an automatic Axis Calibration feature on the Mach3 (Alt-6) Settings page.
Hope this helps,
Tweakie.
The Mach3 manual can be found here;
http://www.machsupport.com/help-learning/product-manuals/