Anwar late Wednesday addressed a rally of supporters dressed in black, to protest Sunday's polls, who filled a stadium and spilled out into surrounding areas, swamping a corner of the capital Kuala Lumpur in hours-long gridlock.
Declaring the elections the "death of democracy", he called for a sustained campaign against electoral bias and cheating that he says props up an "arrogant, racist regime".
"We will go to every corner of this country," Anwar declared, prompting roars from the multi-racial crowd.
"We will continue to struggle and we will never surrender!"
The huge turnout and the charismatic Anwar's call for similar rallies across the country upped the ante in a campaign by the opposition to paint the elections as a fraudulent victory for the regime that has ruled Malaysia for 56 years.
Anwar has said he would soon produce evidence to prove the elections were riddled with fraud by what he calls an "illegitimate" Barisan Nasional (National Front) government headed by Prime Minister Najib Razak.
Najib's government has hotly denied the allegations and urged its opponents to accept the result gracefully.
In the United States, a spokesman for President Barack Obama congratulated Najib on the win but said: "We note concerns regarding reported irregularities in the conduct of the election, and believe it is important that Malaysian authorities address concerns that have been raised."
Spokesman Jay Carney added: "We look forward to the outcome of their investigations."
Najib said before Wednesday's rally that the gathering was "calculated to create unrest".
But the crowd was more festive than angry, waving opposition party flags and sounding vuvuzela horns.
Supporters filled the stadium's 25,000 seats, with at least twice that many on the football pitch. Thousands more clogged surrounding neighbourhoods.
Pictures circulated on social media sites showing Anwar being whisked to the venue on the backs of motorcycles to beat the standstill.
"The government has stolen it. This is not the people's will," said Kenny Lim, 32, an engineer.
"We come out peacefully to fight for a clean election -- that's why we are here."
Police had earlier threatened to arrest participants at Wednesday's rally but with tension high over the country's closest-ever election result, no security presence was seen.
Anwar has battled Barisan since he was ousted from its top ranks in 1998 and jailed for six years on sex and corruption charges widely seen as trumped-up.
He called for another rally in his northern home state of Penang on Saturday, with more to follow around Malaysia.
The opposition has made a host of electoral fraud allegations including suspicious handling of ballot boxes and suspected foreigners being drafted in to cast ballots.
Meanwhile, indelible ink -- meant to thwart multiple voting -- easily washed off.
A report released Wednesday by two independent watchdogs said the polls were marred by "serious flaws" in the electoral system.
The election was "only partially free and not fair", said the report by the Institute for Democracy and Economic Affairs and the Centre for Public Policy Studies.
The vote was touted as the first in which the opposition had a chance to unseat the ruling coalition, which has governed since independence in 1957.
Barisan retained a firm parliamentary majority despite winning less than half the popular vote, a factor blamed on Barisan's tinkering with electoral districts.
The opposition had campaigned on a vow to free up Malaysian society, end what it calls the divisive race politics of the Malay-dominated Barisan, and reform controversial policies that give majority Malays advantages.