Solution
This issue was addressed in CARs 481652 and 413948 and was fixed in NI Vision Development Module 16 as discussed in the
readme.
Code generated for certain algorithms in Vision Assistant did not work the second time the main VI was called if the application disposes all buffers in the main VI.
If using a previous version and you are unable to update, consider using the
IMAQ Dispose VI to dispose the images manually, instead of choosing to dispose all images.