Three out of four police forces are set to take part in the 10-week national sting operation that begins today.
The Home Office has announced that £1.2m in extra funding has been allocated for the test purchase operation and 166 out of the 227 police basic command units (BCUs) across England and Wales are taking part.
It's absolutely critical that pubs focus on delivering the Challenge 21 scheme - the penalties for failing the forthcoming sting operations are extreme British Beer & Pub Association director of communications Mark Hastings.
A Home Office spokesman confirmed that BCUs have been told to target up to 20 licensed premises that they believe may be serving under-18s during the Tackling Underage Sales of Alcohol campaign.
Each venue can be tested up to five times during the campaign, which runs until 15 July.
The spokesman said venues that fail the operation will be told so "as soon as is practically possible".
Police forces are also planning to use tougher new powers to clamp down on problem premises during the campaign.
Under new laws that came into force on 6 April, pubs can be closed for three months - and premises licence holders fined £10,000 - if three underage sales occur within a three-month period.
British Beer & Pub Association director of communications Mark Hastings said: "We've been consistently sending alerts on the need to focus on Challenge 21 (the age-check policy) and companies have consistently been reminding managers and licensees that the campaign is on the way.
"It's absolutely critical that pubs focus on delivering the Challenge 21 scheme in their business and do it rigorously, otherwise the penalties for failing the forthcoming sting operations are extreme."
Bar Entertainment & Dance Association executive director Paul Smith said: "I think if a premises passes the first couple of times, it's a good premises. I don't think they have to go back and back; it seems unnecessary."