Punchline: Yes! Business-to-business applications should have a sense of humor just like consumer applications.
Enterprise users are human beings. They have Facebook accounts and Twitter accounts just like you do. They watch sports and sit-coms on TV just like you do. They laugh, they cry and have emotions just like everyone else. Enterprise users have human psychology just like small business customers. So why do we treat them like soulless robots? Why do we design products that are so serious and have no irreverence or cleverness built into them?
However, being cute or clever can always be taken too far. I have devised the Joke-o-meter to demonstrate this phenomenon.
The important thing is to not go over the line. The line is an important psychological barrier that turns a joke into a problem. The funny-factor diminishes significantly at the line. When someone says, “That’s over the line”, this is what they mean. Of course, some people (like myself) love it when a joke crosses the line a bit too far. See chart below.
The key thing to remember as a designer is that you must maintain a sense of humor about yourself and bake that into the application whenever possible. Don’t go over the line, but take advantage of the fact that your customers are human beings. Figure out what “fun” means to them and deliver.