Opinion polls have suggested big business is broadly in favour of staying in the EU.
British politicians made their final pitch to a bitterly divided electorate on the eve a crucial referendum to persuade undecided voters of the merits of remaining in or leaving the 28-member EU with polls showing a razor-tight race whose outcome could shape Europe's future.
In the biggest backing yet for the "Remain" camp, 1,280 business leaders, which included representatives of 51 FTSE 100 companies, signed a letter warning that Brexit – or Britain's exit from the EU - would mean "economic uncertainty and put jobs at risk".
Their warning came on the last day of the four-month-long campaigning before polling booths open at 7 a.m. local time on Thursday with the final result expected early on Friday.
More than 46 million people are eligible to vote in the referendum in which people are being asked to choose whether the UK should stay in the European Union or leave in the first vote on the UK's links with Europe for more than 40 years.
Opinion polls have suggested that while big business is broadly in favour of staying in the EU, small firms have been evenly split in what looks like a photo-finish with one poll showing "Remain" at 45 per cent and "Leave" 44 per cent, with 11 per cent undecided.
Prime Minister David Cameron welcomed the support from top businesses as he kicked off the final hours of his campaigning, stressing that the UK enjoyed a "special status" within the EU and the "best of both worlds".
Cameron, who has appeared alongside former Prime Minister John Major and former Labour leader Harriet Harman in Bristol, said that the decision will be irreversible and there will no coming back if the UK votes to leave.
"You can't jump out the aeroplane and then clamber back through the cockpit hatch," he told BBC Radio 4's Today.
Leaving the EU would be a "massive problem" for the UK, he said, doing "untold damage" to economic growth, jobs and family finances and hindering the opportunities and life chances of future generations.
Speaking to the BBC, he said "We are not shackled to a corpse. You can see the European economy's recovery. It's the largest single market in the world.
Making a personal plea to those who fear greater European control, he described himself as a "deeply patriotic person".
"We have not been invaded for 1,000 years, we've got institutions that have served us well. I don't want to give that up to some sort of 'United Europe' and that's not what we're going to do.
But Boris Johnson and other Leave campaigners said only a vote to leave the EU could give the UK the freedom it needs to set its own course, rejecting the economic forecasts suggesting the country would face a downturn following Brexit.
The former mayor of London urged people to "believe in our country" and seize the moment.
"This is a crucial time, lots of people will be making up their minds, and I hope very much they will believe in our country, believe in what we can do," Johnson said.
In his closing speech, the UK Independence Party (UKIP) leader Nigel Farage said it had been a "long, lonely road" for him and his party - which has campaigned for EU exit for more than 20 years - and he believed his party's supporters would "crawl over broken glass" to vote for Brexit.
He urged others yet to have made up their mind to "vote with their heart and soul", saying he wanted Britain to be a "normal country that makes its own laws and is in charge of its own destiny in the future".
Labour leader Jeremy Corbyn has sent a message to Labour supporters that "leaving the EU would hurt our economy – which would mean fewer jobs and less money for our NHS and other vital public services".
The political climate leading up to the referendum has been unusually toxic, with both sides accusing the other of lying and making up their arguments.
Last week, the country was shocked by the killing of Labour's Jo Cox, a pro-"Remain" advocate in her first term in Parliament, in her electorate in northern England. She was the first member of Parliament to be killed in office in 26 years.
Brendan Cox, her widower, said that she had worried before her death about politics becoming "too tribal and unthinking."
"She was very worried that the language was coarsening and people were driven to take more extreme positions," he said.
Meanwhile, polling booths up and down the UK are preparing for the big day.
At the close of the polls, thousands of sealed ballot boxes will be collected from schools and church halls which double up as polling stations and transported to one of 382 counting venues across the UK.
In a departure from the norm, no major broadcasters have commissioned any exit polls over concerns about accuracy following the fiasco of the 2015 General Election which had wrongly predicted a hung Parliament.
The result will be declared by Jenny Watson, the chair of the UK's Electoral Commission and the referendum's chief counting officer at Manchester Town Hall on Friday morning.
Thunderstorms and flooding is forecast in many parts of the UK, which could affect turnout.
According to pundits, the "Leave" side is expected to benefit the most from a low turnout.