If Vista recognizes that it can be used as a ReadyBoost device then yes. You can set it back up.
But if you ahve more than 1GB of RAM. ReadyBoost is a waste. Plus ReadyBoost should be double your RAM. So you should have @GB of ReadyBoost for a1GB system. Since 4GB is the Max the Max RAM you should have is 2GB.
Anything mroe. You dont use it anyways.