The ITV period drama, which has previously won Emmys, Baftas and Golden Globes, has never scooped the award for best drama series at the TV Choice Awards, despite being nominated in the same category four years ago.
But the sixth and final series of Downton saw off competition in the category from Happy Valley, Line Of Duty and Peaky Blinders.
Dame Barbara Windsor was honoured for her outstanding contribution to television at the awards, now in their 20th year.
Declaring herself “absolutely delighted” with the gong, the veteran actress said: “It is such an honour to be recognised by TV Choice and it had not been something I ever expected to receive so I am extremely grateful.
“I have been in the business for over 66 years and I have been fortunate to have been involved in some truly outstanding productions that all mean so much to me, so to know that they also mean a lot to the audience is extremely touching.
“Everyone knows how much love I have for EastEnders and that show has played such a pivotal part in my career, without it I may not be receiving this award. Although Peggy may no longer be with us, she and the show will never leave my heart.”
Walford stalwart Peggy, who had been played by Dame Barbara since 1994, took her own life after being diagnosed with terminal cancer in scenes broadcast in May.
Elsewhere, the best actor award went to Tom Hiddleston for BBC One’s The Night Manager. He beat Doctor Who’s Peter Capaldi, Cillian Murphy for Peaky Blinders and Jim Carter for Downton Abbey.
Happy Valley’s leading lady, Sarah Lancashire, saw off the likes of Downton Abbey’s Laura Carmichael and Peaky Blinders star Helen McCrory to be named best actress.
ITV soap Emmerdale was the other big winner on the night. The long-running series was named best soap over EastEnders, Coronation Street and Hollyoaks for the first time.
Doctor Foster was named best new drama, with The Great British Bake Off winning best talent show ahead of Strictly Come Dancing, The X Factor and Britain’s Got Talent.
The ITV series I’m A Celebrity... Get Me Out of Here!, presented by Ant and Dec, took the best reality show prize, the first of two awards won by the Geordie duo on the evening. They also walked away with best entertainment show for Ant & Dec’s Saturday Night Takeaway.
It was a night of success for ITV, which claimed eight of 14 main awards, including best daytime show for This Morning and best comedy for Birds Of A Feather. Channel Four took two awards, with Gogglebox winning best lifestyle show and Sunday Brunch named best food show.
The star-studded ceremony, hosted by comedian Jo Brand, was held at The Dorchester in London.