Karzai criticised the decision by all 15 remaining candidates to withdraw from the presidential race on Saturday saying they must respect the will of the people.
"Just because 15 people have said 'No', we can't deny the votes of millions," Karzai told a news conference after all his rivals announced their decision.
"It's too late in the day for a boycott. Millions have voted in the rain, the snow and the dust storm and we should respect their decision," he said.
Afghanistan's presidential elections were plunged into crisis after Karzai's rivals claimed irregularities.
An aide to Yunis Qanuni, the main challenger to President Karzai, said the opposition candidate was demanding a suspension of the first Afghan exercise in democracy for decades to prevent further fraud.
Another challenger, Abd al-Satar Sirat, said candidates called for a halt after an early morning interruption when it appeared the ink meant to prevent multiple voting could be rubbed out.
"Today's election is not a legitimate election. We are not a part of today's election," Sirat said after an emergency meeting on Saturday.
But the election commission said the setback was a case of inexperienced electoral workers in Kabul polling stations using the wrong ink and had quickly been rectified.
"Halting the vote at this time is unjustified," Ray Kennedy, a member of the Joint Election Management Body, told reporters. "Afghans have turned out in large numbers [to vote] and the process has been safe and orderly."
Kennedy was speaking just minutes before polling stations officially closed at the appointed time of 4pm (1130 GMT) and hours after all the candidates arrayed against Karzai said they were boycotting the election because of problems with systems put in place to prevent illegal multiple polling.
The move by 15 of the 18 candidates in the fray effectively leaves Karzai as the only one running - two others withdrew in his favour earlier this week.
Qanuni's aide said the boycott call had also been prompted by claims that while polling stations were open in areas noted for Karzai supporters, they had closed in areas popular with opposition supporters.
Voters had flocked to polling stations as soon as they opened at 7am despite threats of violence from supporters of the former Taliban and al-Qaida.
US-backed Hamid Karzai was
chosen as interim leader in 2002
Karzai, chosen as interim leader by a council of tribal leaders in June 2002 and the US favourite, only managed one campaign rally. His first attempt at rallying the people was aborted by an assassination attempt.
The urbane 46-year-old is running against 17 other candidates, including one woman, a French-speaking poet and two regional commanders.
Chief rival Qanuni is the favourite of the powerful anti-Taliban ethnic Tajik clique from the northern Panjshir Valley.
With few policies in the campaign, most Afghans were expected to vote along ethnic lines, giving Karzai a strong chance as he hails from the southern Pashtun majority, shared by the former Taliban rulers.
Tajiks, Uzbeks, Hazaras and Baluchis make up the rest of the mosaic of Afghanistan's tribes.
Fears were high before the poll that regional commanders, who still wield power through their private armies, would intimidate voters.
Most candidates were expected
to vote along ethnic lines
The still-undersized Afghan army and police force are inadequate to guard all 5000 polling booths, so militiamen from regional commanders' private armies have been deployed in many areas.
Only 230 international monitors are braving threats of Taliban violence, which has claimed 12 electoral workers' lives since May. Almost 4000 local observers are assisting.
The country was reported to be largely peaceful except for a firefight between US-led troops and armed fighters south of Kabul.
Three soldiers from the US-led force were wounded, Major Scott Nelson said.
Concerns were raised briefly on Saturday when a western official reported a bombing at a polling station in the north, but Afghan officials and international peacekeepers said no such blast had occurred. They said a truck fire had broken out, but caused no injuries.
A flurry of rockets landed in several cities around the country on Thursday and Friday, including one that hit a parking lot near the US embassy and another that injured a young girl and an old man in the eastern city of Jalalabad.