Last season's Challenge Cup saw the Cherry and Whites beaten by Newport Gwent Dragons on home soil at exactly this stage. It's a result Humphreys is hoping won't be repeated.
He said: "For a lot of us the Dragons last year was a particular low point so we know how important this game is for us in the context of the season.
"We're conscious we need to build on last week's performance at Bristol, which was good, but we obviously need to be better this week to get the result. Our mind's very much on the threats the Blues have across their team with some of their players coming back from the Six Nations.
"We know if we give them space and time on the ball they've got players who will hurt us. We defended very well last week and we brought some real energy and line speed to our defence - that's something we've got to bring again."
Gloucester finished top of Pool 1 with 25 points, but are currently without injured skipper Greig Laidlaw. Hooker Richard Hibbard has been in fine recent form for the English side, and will be looking to continue his impressive run against the team from his homeland.
Humphreys added: "Being back at Kingsholm, coming off what was a disappointing result against Harlequins and all the implications of that result, we want to show the supporters what this team is about. We want to build on last week's win and bring some pride again with a big performance.
"We've also got to be better with the ball. That's a big part of our focus, that we get the basics right, we get our set piece right, and when we get the ball we hold on to it."
Cardiff Blues are set to welcome back Wales flanker Sam Warburton for their short trip across the border, as they look to get back to winning ways following defeat at Leinster.
"This is why you play, to have the opportunity to chase cups like this," said wing Blaine Scully.
"This is now everything to us. It is where all our focus is. It will be pretty big, our biggest game of the year and a real raw opportunity for us to go out make a statement and put in a performance.
"Kingsholm is pretty unique and one of the great grounds. They obviously love everything about their rugby team, it's an energetic crowd and can be hostile but as a competitor that's a great place to go and compete.
"It's fun to be in that environment and feels like it's you against the world. You have to embrace that moment and enjoy it for what it - an amazing opportunity to play in a great ground, against a great opponent in a really meaningful game."