The campaign, created by TDA, aims to inspire people to support the charity with a monthly direct debit which will help fund a puppy's training to become a qualified guide dog.
The ad shows an everyday journey that a qualified guide dog and its blind or partially sighted owner might make via public transport, including negotiating steep staircases, crowds, roadworks and cyclists.
Footage of the fully trained guide dog is interspersed with that of a puppy in training.
Amanda Mitchell-Francombe, head of individual giving at Guide Dogs, said: "Guide dog puppies undergo extensive training before they are ready to provide a truly life-changing level of independence and freedom to a person with sight loss.
"By sponsoring a puppy people can help to fund the puppy's training from six weeks to around 20 months of age, when the young dog will qualify as a working guide dog with its new blind or partially sighted owner."
The ad will air on satellite and terrestrial channels including Sky and UKTV from September 15. Viewers will be encouraged to call Guide Dogs or go online to pledge £1 a week to become a Sponsor a Puppy supporter.