The result, declared early on Friday, showed Labour's John Kane as the winner of a seat previously held by SNP stalwart Councillor Allison Hunter, who died recently.
Mr Kane, a full-time carer for his disabled partner who has been resident in the ward for 25 years, beat the the SNP's Helen Walker and 12 other candidates.
The poll was run on the Single Transferable Vote system. Turn-out was 20.06%.
Mr Kane's victory, on a 7% swing, strengthens Labour's grip on Glasgow, which had been a key target for the SNP at the 2012 elections.
It also leaves Govan as the only electoral ward in the city without any Nationalist representation. Given that the area lies on the doorsteps of Deputy First Minister Nicola Sturgeon, along with Scottish Labour leader Johann Lamont and her deputy Anas Sarwar, the outcome of Thursday's poll will be seen by some as an electoral barometer 11 months before the independence referendum.
Councillor Gordon Matheson, Labour leader of the council said: "There's no way the SNP can spin their way out of this dramatic result. What odds would the bookies have given a year ago on Govan ward being SNP-free?
"With less than 12 months until the referendum, support for the nationalists has slumped in Glasgow. Councillor John Kane joins a strong Labour administration which continues to focus on the issues that matter to people, not the abstraction of separatism."
Labour has suffered two historic parliamentary by-election defeats in Govan to the SNP. Yesterday's result was the latest of a disappointing run of results for nationalists since last October's announcement of the referendum.
The SNP's only comfort was that its vote share was down by just 1.5% but this was probably due to the absence this time of several high-scoring minor party runners from 2012.
John Flanagan, standing under the banner of No Bedroom Tax - No Welfare Cuts came third in the field of 14 candidates.
If the movement in Glasgow was to be repeated at Dunfermline on October 24, Labour would win the former SNP Holyrood seat held by Bill Walker in a landslide.
Some voters will have two ballot papers on October 24 as there is a Fife Council by-election in Dunfermline South ward on the same day as the Holyrood contest.
Meanwhile, the Conservatives have held their seat in the Tweeddale West by-election.
Keith Cockburn was elected to replace Nathaniel Buckingham, who resigned in July.
Councillor Cockburn will join councillors Willie Archibald (SNP) and Catriona Bhatia (Liberal Democrat) to represent the Tweeddale West ward.
The overall turnout for the ward was 35.4%.
Returning Officer, Tracey Logan said: "Thanks to the hard work and commitment of our staff the by-election has run very efficiently. I welcome Councillor Cockburn, and council officers and elected members alike look forward to working with him in the future."
RESULT: Glasgow City - Govan: First count Lab 2055, SNP 1424, No Bedroom Tax 446, C 215, Ukip 113, Green 112, Ind 103, Lib Dem 73, Christian Party 60, Ind 52, Communist Party of Britain 35, Solidarity 28, Britannica Party 19, Scottish Democratic Alliance 1. Lab gain from SNP. Swing 6.8% SNP to Lab.