The three-week trial on the 23.4m long 'up' escalator at Holborn station is taking place during peak hours.
Transport for London said the move will "increase the capacity" and prevent "queuing at the bottom" as people stand on both sides of the escalator.
More than 56 million passengers use the station every year.
Usually passengers stand to the right, leaving the left side free for people to walk up or down the escalators.
Holborn Tube station, one of the busiest on the network, is set to get a new entrance and an enlarged ticket hall in order to increase its capacity by 80% by 2022.
Peter McNaught, operations director for the Central line, said: "In the meantime, as part of a three-week trial to reduce queues and keep people moving through the station, we are encouraging customers to stand on both sides of the 'up' escalators during busy periods.
"We believe this will increase the capacity of the escalators as more people will be able to board, preventing queuing at the bottom of the escalator."

The lawsuit filed in a federal court in San Francisco argues that the policy is unconstitutional.
Women can serve in front line positions in the US military, but they are barred from ground combat units.
A Pentagon spokesman said 14,500 combat positions had been opened to women under the current defence secretary.
He added that Defence Secretary Leon Panetta had "directed the services to explore the possibility of opening additional roles for women in the military".
But ACLU lawyer Ariela Migdal said the changes so far were "not enough". The lawsuit argues that continuing restrictions violate servicewomen's constitutional rights to equality.
One of the plaintiffs, Marine Corps Capt Zoe Bedell, said existing rules had blocked her advancement in the Marines.
"The military is the last place where you are allowed to be discriminated against because of your gender," she said.
Women account for about 14% of the 1.4 million active US military personnel.
The lawsuit launched on Tuesday says they are barred from 238,000 positions, but also alleges that they are already serving unofficially in combat units.

Russell Goodway told BBC Radio Wales that councillors and officials were the "experts" and should "seriously develop" a set of proposals.
Former Public Services Minister Leighton Andrews had been planning to cut the 22 councils to eight or nine.
But First Minister Carwyn Jones has said the Welsh Government will have to rethink its plans to win wider support.
Mr Andrews is no longer in government after losing his Rhondda seat at the election in May, and the Labour manifesto did not include a commitment on the number of councils it would like to see.
As a minority government, Labour will need the support of other parties if any cut is to happen.
Speaking on the Good Morning Wales programme on Monday, Mr Goodway said: "The status quo isn't an option.
"You really cannot maintain 22 local authorities unless you're prepared to dismantle local government as we know it and create other organisations or bodies that would be responsible for some of the major services such as education, social services and possibly economic development.
"I would like to see the experts in the field, who happen to be those that run local authorities - whether they're councillors or council officials - coming forward with well-developed proposals to meet the government's aims.
"I do not want to see a top-down reorganisation imposed on Welsh local government, particularly at a time when you're seeing a lot of new faces in the assembly, people who don't have the expertise in local government to shape those services.
"I really do hope now the Welsh Local Government Association and the leaders in local government will come forward and seriously develop a set of proposals, hopefully capturing the city region dimension."
On Sunday, the first minister told BBC Radio Wales he wanted to talk to opposition parties about the future of local government.
"Clearly, the map we published before the election, that is not going to gain support across the assembly, that's obvious," Mr Jones told the Sunday Supplement programme.
Meanwhile trade unions representing council workers have asked for urgent talks on the issue with the new finance and local government secretary Mark Drakeford.
Dominic MacAskill, head of local government in Wales for Unison, said: "We are in a state of limbo and, in that state of limbo, the workforce is anxious and fearful of the future."
Commuters are being asked to stand rather than walk on the left side of escalators at a central London Tube station to see if it can cut queues.
[NEXT_CONCEPT]
The American Civil Liberties Union has filed a challenge on behalf of four US servicewomen against a ban on women being deployed in most combat roles.
[NEXT_CONCEPT]
Councils should draw up fresh plans to reorganise local government, a former leader of Cardiff council has said.