Proof of double vaccination will now be required to enter nightclubs and large events, either using a paper certificate or the NHS Scotland Covid Status App.
The scheme was launched on October 1, but the Scottish Government allowed an initial grace period during which it was not legally enforced.
Health secretary Humza Yousaf said the government has listened to feedback from businesses, and they should have had time to prepare themselves for enforcement.
But the plan remains controversial, with opposition parties still calling it “unworkable”.
The initial launch of the app was plagued by download issues, which the government said were due to the high volume of people attempting to access it at the same time.
The Scottish Conservatives have called repeatedly for the programme to be delayed.
Shadow Finance and Economy Secretary Liz Smith said businesses had not had enough time to prepare.
She said: "Scottish businesses are barely more prepared today than they were a fortnight ago when this chaotic scheme was rushed in.
"We have repeatedly called for delays and for significant practical changes to be made, so that businesses and sporting facilities have sufficient time to prepare for these burdensome checks.
"They will have no choice but to fully comply with this unworkable scheme and will be forced ever closer to a cliff edge.
"This has the potential to seriously undermine Scotland's economic recovery in the run-up to the holiday season."
Scottish Labour described the certification scheme as a “mess”.
“The shambles that followed the release of the vaccine passport app was a predictable disaster,” said health spokesperson Jackie Baillie.
“Weeks have passed and we have seen no change of plans despite concerns from the public, public health experts and businesses.”
The Scottish Night Time Industries Association, which represents nightclubs, launched an unsuccessful legal challenge against the programme.
The group declined to comment on nightclubs’ state of readiness ahead of the enforcement date.
Celtic has confirmed that vaccine passports will be needed by fans attending the Europa League match against Ferencvaros on Tuesday.
Certificates will be required at all unseated indoor live events with more than 500 attendees, and unseated outdoor events with more than 4,000.
They will also be needed for entry to nightclubs, defined as any venues open after midnight featuring alcohol, music and dancing.
Under 18s are exempt, along with anyone who cannot be vaccinated for medical reasons, people who took part in vaccine trials and those working or performing at the venues.
Mr Yousaf said: "This is a very limited scheme and we hope this will allow businesses to remain open and prevent any further restrictions as we head into autumn and winter.
"This virus has not gone away and vaccine certification will have a role to play in keeping transmission under control as part of a wider package of measures. It adds a further layer of protection in certain higher risk settings.”
A message from the Editor:
Thank you for reading this article. We're more reliant on your support than ever as the shift in consumer habits brought about by coronavirus impacts our advertisers.
If you haven't already, please consider supporting our trusted, fact-checked journalism by taking out a digital subscription.