Medecins Sans Frontieres says hospitals it supports in Syria treated about 3,600 patients with "neurotoxic symptoms", of whom 355 have died.
It said the patients had arrived in three hospitals in the Damascus governorate on 21 August - when opposition activists say chemical attacks were launched against rebels.
But MSF says it cannot "scientifically confirm" the use of chemical weapons.
Both sides in the conflict accuse each other of using chemical weapons.
MSF says staff at the hospitals described a large number of patients arriving in the space of less than three hours with symptoms including convulsions, extreme salivation, contracted pupils and sight and respiratory problems.
The charity said many were treated with atropine, a drug administered to those with "neurotoxic symptoms".
"MSF can neither scientifically confirm the cause of these symptoms nor establish who is responsible for the attack," said MSF Director of Operations Bart Janssens.
"However, the reported symptoms of the patients, in addition to the epidemiological pattern of the events, characterised by the massive influx of patients in a short period of time, the origin of the patients, and the contamination of medical and first aid workers, strongly indicate mass exposure to a neurotoxic agent.
"This would constitute a violation of international humanitarian law, which absolutely prohibits the use of chemical and biological weapons."
MSF's disclosure came hours after the UN disarmament chief Angela Kane arrived in Damascus to press the Syrian government to allow access to the site of the alleged chemical weapons attack.
France has joined the UK in accusing Bashar al-Assad's forces of carrying out the attack in the capital's eastern suburbs on Wednesday.
US President Obama has said he is weighing his options and described it as a "big event of grave concern". More