The left eye was clearly having a growing impact on him, though, and he was eventually forced to take a knee in the 11th round while under pressure from Spence.
Brook and his promoter Eddie Hearn confirmed that a broken left eye socket was suffered.
"He's a true champion and I wanted to fight him", said Spence, who gained his ninth straight stoppage victory.
With the hometown crowd backing him all the way, Brook bravely fought on like the warrior he is but went down in the tenth and finally succumbed to the severity of his injury in the eleventh.
Friday's weigh-in represented Brook's first return to the welterweight division since gaining 13lbs to challenge Gennady Golovkin last September in his most recent fight.
"I'm gutted, devastated that in front of my own fans I've lost my belt".
"I watched some of his fights and he likes to fight at a certain pace". It's also rare to see a challenger who's willing to jump into the lion's den and come out victorious.
Whilst Spence boasts a flawless record - winning all of his 21 professional fights, with 18 KOs.
Spence, 27, of DeSoto, Texas, appeared relaxed before the fight while Brook, born and raised in Sheffield, looked a little tense.
In his fourth title defence, Brook quickly discovered the hype surrounding Spence is more than justified as the 27-year-old made a fine start.
'I tried to get through it but kept getting double vision and then it would come back into line but in the 10th and 11th round it stuck there and that's why I went down on one knee'.
Undefeated #1 contender Errol "The Truth" Spence, Jr.
But Chris Eubank Jr wasn't as charitable: "How do you just give up your world title with only 2 rounds to go because of a swollen eye?" "I knew from round seven that the eye had gone and progressively as the rounds went on". I can face adversity and I can win.
Those watching the action rightly commended the Sheffield man for his wherewithal, and paid tribute to a special prospect in Errol Spence Jr., who now moves to 23-0 in his boxing career. "I got on top of my weight early in camp so I came here fit and healthy and firing on all cylinders".