Britain's foreign minister Boris Johnson said Friday it was "overwhelmingly likely" President Vladimir Putin directly ordered the attack.
Russian Federation also said it was halting the activities of the British Council, Britain's global organization for cultural relations and educational opportunities, across the country.
"If further actions of an unfriendly nature are taken against Russia, the Russian side reserves the right to take other retaliatory measures", the ministry said.
The Russian Foreign Ministry summoned the British ambassador, Laurie Bristow, to a meeting on Saturday morning in central Moscow at headquarters during which he was informed of the measures.
"We will always do what is necessary to defend ourselves, our allies and our values against an attack of this sort, which is an attack not only on the United Kingdom, but upon the global rules-based system on which all countries, including Russian Federation, depend for their safety and security", Bristow told reporters after being informed of the expulsions.
Both victims remain critically ill in hospital after being poisoned with what authorities believe was the military-grade nerve agent Novichok.
The Russian Foreign Ministry also said in a statement that it is ordering the closure of the British Council, a government organization for cultural and scientific cooperation, and that it is ending an agreement to reopen the British consulate in St. Petersburg.
Britain, the United States, Germany and France have jointly called on Russia to explain the attack, while US President Donald Trump has said it looks as if the Russians were behind it.
Police officers at the scene of the nerve agent attack in Salisbury, England..
British police appealed Saturday for witnesses who can help investigators reconstruct the Skripals' movements in the crucial hours before they were found unconscious.
She tells British lawmakers that the military-grade nerve agent used was identified as belonging to a group of nerve agents known as Novichok and of a type developed by Russian Federation.
By releasing the image of Mr Skripal's vehicle, police are hoping to establish more details about the movements of the Skripals on the morning of the attack.
A post-mortem examination found the 68-year-old died from "compression to the neck".
The council was granted a Royal Charter in 1940, which set out its mission as "promoting a wider knowledge of the United Kingdom and the English language overseas and developing closer cultural relations between the United Kingdom and other countries".
Russia's Investigative Committee said it had launched its own criminal proceedings in connection with the "attempted murder of a Russian citizen, Yulia Skripal" in Salisbury and the "murder" of Glushkov in London. After several days of investigation, British authorities determined that they were poisoned with a nerve agent known as Novichok, which is believed to be unique to Russian Federation. The Kremlin has always denied the accusation.
"The onus remains on the Russian state to account for their actions and to comply with their worldwide obligations".
Skripal is a former colonel in Russian military intelligence who was jailed in his country for betraying agents to Britain's MI6 secret service and moved there in a spy swap in 2010. He was granted refuge in the UK after a high-profile spy exchange between the United States and Russian Federation in 2010. The way to make the point stick in Moscow is to start cutting off access to funds and markets, and let the Russian economy in an oil-glut world do the rest.