The last Melbourne-to-Sydney XPT train arrived at Sydney's Central Station at 7am local time this morning, while the last Sydney-to-Melbourne train left yesterday evening.
The 75 people who arrived in Sydney on the last train were brought out carriage by carriage, and walked into a cordoned off area to have their temperatures checked and details taken for contact tracing.
It comes after Nationals Senator Bridget McKenzie said she is concerned regional communities will be hit the hardest by the closure decision.
NSW has closed its border to anyone from metro Melbourne as of midnight last night, and after 11.59pm tonight that travel ban will be extended to all Victorians.
With South Australia also having aborted plans to re-open its borders, from midnight tonight's Victoria's population of 6.3 million will be locked in.
There will be exemptions for residents in border communities who will still need to cross state lines for work or health reasons.
But Mackenzie told Sky News there was considerable "anger and confusion" over the announcement.
"We have a community of 100,000 people between Albury and Wodonga," she said.
"We share a workforce, we share education system and a health system – for instance you can only have babies here in Wodonga – and it begs the question, where does a new mother go when she heads back to Albury, does the father have to quarantine?"
"There's a lot of confusion and anger around a community that really sees itself as two cities, one economy, and one community," she added.
Senator Mackenzie said that since the outbreak was confined to Melbourne, it would have made more sense to lock down the city and leave the regions alone.
Victorian Premier Daniel Andrews said the decision to close borders between Australia's largest states followed a phone call with Prime Minister Scott Morrison and NSW Premier Gladys Berejiklian.
It came after Victoria recorded a record 127 cases yesterday, including 16 in the public housing towers that have been locked down.
People who live on the border will be eligible to receive a special permit to travel across the border for work.
It's the first time the border between the two states has been closed in 100 years, showing the level of anxiety about the potential spread of the virus from Victoria into NSW.
The closure will be enforced by NSW authorities to avoid draining Victorian resources committed to fighting the surge of cases across the state.
Police forces said drone technology would be used to assist in implementing the closure.
Victoria enacted a "hard lockdown" on nine public housing towers in Melbourne – effectively confining 3000 people to their homes – after a cluster of cases were found.
Residents in at least 36 suburbs deemed to be "hot spots" were put into a separate lockdown, but can still leave home for work, school, exercise and essential shopping.
NSW Police Commissioner Mick Fuller said police would monitor the 55 border crossings between the two states, with 400 police to be deployed and barriers set up at popular crossing points.
He warned there could be major delays for those attempting to cross the border before midnight tonight.
"We know there are four primary road crossings, 33 bridges, two waterway crossings and multiple smaller roads," he said.
"The task is not lost on me in terms of the enormity of the logistics of this operation. There are 11 local government areas and five police districts, which will all have to pull together to make sure this operation is a success."