Brooke Cotchin's visit was one of several breaches announced by the league on Friday with $185,000 fines being handed out to the Tigers, Kangaroos, Blues and Hawks.
$25,000 of the $45,000 fine handed down to the Tigers is suspended, pending any second breach.
She captioned the since expired post: "The most incredible experience today…. My skin was in desperate need of some love. I knew I was in the right place as soon as I walked through the doors of Esteem."
Collingwood president Eddie McGuire said on Friday night it was unfair for the AFL to punish clubs for the actions of players' wives.
"It's an interesting one. The club's have got to be involved, but it has to fall on the individual," he said.
"There's not much (Richmond chairwoman) Peggy O'Neil in Melbourne can do to stop Trent Cotchin's wife from going to a day spa. The fact that she put it up on Instagram shows she wasn't trying to hide it. It was just a misunderstanding."
AFL General Counsel Andrew Dillon released a statement on Friday announcing a second breach of the protocols will result in a fine of up to $75,000 in addition to the added $25,000 from the suspended first offence fine.
A third breach could result in clubs being stripped of premiership points.
"Our primary focus, like everyone around the country, is to play our role in continuing to keep safe the communities in which we play. We know it is a privilege to be able to continue our competition which provides jobs for thousands of people around the country, and joy to millions of fans." Dillon said.
"The protocols are in place to not only protect players, officials, staff, and now their visiting families, but also the wider community in which we have been given the opportunity to train and play.
"We all have to modify our behaviours in order for the competition to continue in a safe manner."
It was earlier revealed Hawthorn had received a fine of $50,000 for players breaching the protocols during a trip to a snack vendor inside the SCG last week.
Hawks players — who were not selected last week — went to get some chips from a vendor inside the SCG during their loss to the Swans in Sydney.
Footy reporters Sam Edmund, Jon Ralph and Tom Morris all tweeted the news that the Hawks players broke the protocols despite being escorted by security when they went to find some chips.