Could not load file or assembly or one of its dependencies. Access is denied. The issue is random, but after it happens once, it continues

For my scenario, I found that there was a identity node in the web.config file.

<identity impersonate="true" userName="blah" password="blah">

When I removed the userName and password parameters from node, it started working.

Another option might be that you need to make sure that the specified userName has access to work with those "Temporary ASP.NET Files" folders found in the various C:\Windows\Microsoft.NET\Framework{version} folders.

Hoping this helps someone else out!

Had the same issue, fixed with setting the parameter "Enable 32-bit applications" to "true" (in advanced settings of iis application pool).

My solution is as follows:

I didn't find a root folder under C:\Windows\Microsoft.NET\Framework\v2.0.50727\Temporary ASP.NET Files.

Google told me that it might be a permission issue against current user, then I found I have a current Identity: IIS APPPOOL in the malfunctioning server where the rest of the server has Current Identity: NT AUTHORITY\NETWORK SERVICE.

Then I changed Current Identity from IIS APPPOOL to NT AUTHORITY\NETWORK SERVICE.

From here, I found that resetting the web app rebuilds the temporary ASP.NET cache, solving the issue.