Chuck's spy will refers to Charles Bartowski's testament, a precaution that almost every government field agent takes in case of their death on a mission. A spy will is usually contained in a small circular box opened by a stick like key. The spy will goes to the spy's family and/or friends and/or loved other ones in the event of their death.
Chuck is seen making out a spy will at the end of an episode after Sarah gave him hers. The contents are unspecified, except as follows; "My name is Chuck Bartowski, and if you're reading this, than it means I'm already dead. I don't know what will end up killing me, but I have chosen to be a spy and there's consequences to that.
It may be my emotions that end up doing me in, or a slip up trying to protect my friends, or my family, who never wanted me to be a part of this; or it could be the thing I never saw coming, but I know it's coming. "