In second place is the Home Office and Barclays comes third. MI5 secures a position on the Top 100 for the first time, and is one of 25 new entries.
The 2012 Index introducing new weighting for global employers that recognises worldwide support for LGB equality.
Accenture won the award for Employee Network Group of the Year and the Department of Energy and Climate Change is named Most Improved Employer.
Bill Payne, former chief executive of Metropolitan Housing Partnership, was named Individual Champion of the Year.
The results were announced last night at a ceremony hosted in London by Ernst & Young where the Top Employer award was presented by minister for equalities, Lynne Featherstone.
Ben Summerskill, Stonewall chief executive, said: "Competition for a place in the Top 100 was fiercer than ever this year. With new, more demanding criteria, every employer securing a position in the Top 100 has performed impressively - and the participating employers collectively employ over 1.9 million people. The Index remains a powerful tool used by Britain's 1.7 million gay employees and 150,000 gay university students to decide where to take their talent and skills."
Liz Bingham, managing partner for people at Ernst & Young, added: "We believe that a strong commitment to diversity and inclusiveness is not only important for our people, but is also good business in an increasingly competitive and interconnected world."
Jonathan Evans, director general of MI5, said: "The Security Service has worked hard in recent years to promote equality and diversity across all areas of its work. We are pleased to be recognised by Stonewall, but there is still more we can do. We will continue to support lesbian, gay and bisexual staff to make MI5 a truly inclusive place to work."
The Index is based on a range of key indicators which this year included a confidential survey of lesbian, gay and bisexual employees, with over 7,500 participants. This consistently confirmed that the satisfaction levels of gay staff were highest at the top-ranking organisations in the Index.
