Around 100 emergency services personnel will work with Highways England traffic officers in response to the staged crash on Saturday night.
Exercise Dark Knight will be held on the M62 near Eccles, Greater Manchester, at a time when the motorway was already due to be closed for upgrade work.
Around 50 volunteers will take on the role of injured drivers and passengers to bring the incident to life.
This includes 15 members of a local amateur dramatic society painted in fake blood.
Chris Chadwick, North West resilience co-ordinator at Highways England, said: "Our motorways are among the safest in the world and we help to keep them safe by carefully planning what we'd do on the rare occasions when there's a major incident.
"We normally have to do this away from the motorway to avoid disruption to drivers but will be able to make our latest scenario as realistic as possible by carrying out Exercise Dark Knight on the M62 when it's already closed overnight for major construction work.
"We're working closely with the emergency services on the exercise and are grateful to the volunteers who are giving up their time to help make our motorways safer for everyone."
The motorway will be closed from junction 10 to 12 between 9pm on Saturday and 9am on Sunday.