This year's jury sees former Royal Academy Exhibitions Secretary Sir Norman Rosenthal joined by contemporary artists Goshka Macuga and Gary Hume, Liverpool-born artist Ged Quinn and painter Alison Watt.
Previous winners have included David Hockney, Peter Doig and Michael Raedecker.
Judging will take place throughout 2010 and will result in an exhibition at the Walker Art Gallery from September 18 to January 3 2011, forming a central part of the Liverpool Biennial.
Reyahn King, director of art galleries at National Museums Liverpool, said: "The John Moores Contemporary Painting Prize is a unique opportunity for artists to show their work in a major gallery."
Littlewoods founder Sir John Moores, a keen painter, started the exhibition in 1957 and over the last 50 years almost 2,000 artists have exhibited.
More than 100 people have sat on the jury including singer Jarvis Cocker, writer Germaine Greer and artists Peter Blake, Tracey Emin and Jake and Dinos Chapman.
Artists have until February 19 to register, with online registration on www.liverpoolmuseums.org.uk/johnmoores - and images must be sent in by March 5.
Entries have to be original, new or recent paintings within a set size, designed to hang on walls, by someone who lives, or is based, in the United Kingdom.
Full conditions are on the website.