"Design Life"

Here is what the Ansys Help says about Fatigue Damage...

"Fatigue damage is defined as the design life divided by the available life. The default design life may be set through the Options dialog box. A damage of greater than 1 indicates the part will fail from fatigue before the design life is reached."

You decide on the Design Life for your application. That may be a specification on the design, if not, the engineer, product manager or customer must choose a value.

"What is the right load for fatigue testing?"

Are you asking about fatigue testing to obtain material data to create an SN-Curve in Engineering Data?  Here is my answer:

Look at the SN fatigue data for Aluminum Alloy in the General Materials database that comes with Ansys. There are 8 points on this curve for fully reversed loading (R-Ratio = -1).

There are 6 points on the curve for zero-based loading (R-Ratio = 0).

There are also two other R-Ratios with data in them that make up 10 more load sets, for a total of 24 load sets.

Each load set may have had 10 samples that were run on a fatigue testing machine to measure the cycles at failure, so a total of 240 tests. This is the kind of effort it takes to obtain data that can be used to predict fatigue in parts that you design that may be subject to a variety of load cases.

"What is the right load to apply?"

Are you asking about a specific part shape to do fatigue analysis (not testing) and you already have the SN Curve for the material?

Are you asking what pressure to apply to your part shape to obain these values of stress?  That depends on the shape of your part. Note that if your part is long and slender like a tensile test coupon, it may buckle under compressive loads.

What component of stress is used to measure Alternating Stress?

Alternating stress is the difference between the maximum and minimum normal (axial) stresses experienced by a material during each cycle of loading.