The six month trial of standing-only escalators at Holborn station will start on Monday.
During the trial commuters will have to stand on both sides of the escalator, rather than walking on one side.
The scheme was first announced last month in order to boost capacity at one of London's busiest Tube stations.
The thinking behind the counterintuitive move is that the majority of travellers stand on the escalator at Holborn anyway.
This leaves the left, where people usually walk or run, empty. By opening this up to standing passengers, Transport for London will see if it eases the congestion, overcrowding, and number of people queuing to get out.
A previous test held in November and December 2015 found that as many as 30 per cent more customers could use an escalator in the busiest parts of the day if they stood on both sides.
Peter McNaught, Operations Director at London Underground, said in March: "It may not seem right that you can go quicker by standing still, but our experiments at Holborn have proved that it can be true. This new pilot will help us find out if we can influence customers to stand on both sides in the long term, using just signage and information.
"Anyone who wants to walk on the other escalators will be free to do so, but we hope that with record numbers using the Tube, customers will enjoy being part of this experiment to find the most efficient ways of getting around."