Reading is the most important.
The second seems to me more natural. It seems wrong to use an equal sign and then add to it Not.
NotIMHO is best used with boolean functions such as IsNumeric(...)that lends itself to more natural reading of code, for example,
If Not IsNumeric(myNumberString) Then
Not to = False . , ...
If bFlag1 Then
If Not bFlag2 Then
... , ...
If bFlag1 = True Then ' <-- (This is totally redundant) = True
If bFlag2 = False Then
If a <> b = True Then ' <-- please don't ever do this