The Google-owned site has always allowed partners to sell their own ads in and around video clips, but until now has drawn the line at third-party ad-serving.
Yesterday, however, YouTube begun testing FreeWheel, a video-ad serving platform that will enable content providers to serve ads, meaning the same ad could appear in the same content, whether on YouTube or elsewhere on the web.
The move allows YouTube to be part of a greater network of distributors also using FreeWheel, including media giants CBS and Warner Bros, as well as online-only content companies such as Demand Media and Next New Networks.
This is likely to make the site significantly more attractive to big-brand advertisers looking to run far-reaching campaigns across a number of high-traffic video sites.
YouTube has also recently allowed content providers including Sony Pictures and ESPN to place their own video players on the site, enabling them to insert their own ads and attribute the traffic to their own networks.
Google is working hard to boost the amount of ad revenue it generates from YouTube in an effort to cover the huge cost of running the site.
Despite its massive overheads, YouTube is much closer to achieving profitability than previously thought, according to research from RampRamp.
The figures show that the Google-owned video sharing site will lose a mere $174 million (£197m) this year, much less than the $472m (£321m) loss estimated by Credit Suisse.