The South Yorkshire club said the 22-year-old defender is now suspended whilst the investigation takes place.
A positive test for a banned substance which is on the World Anti-Doping Agency’s prohibited list can lead to a four-year suspension.
A statement from the club on Wednesday afternoon read: “Barnsley Football Club have received notification from the FA that they are investigating a potential anti-doping violation by our player, Bambo Diaby.
“The player and club are co-operating and responding directly with the FA. The player is now subject to a playing suspension whilst this matter is under investigation.
“During these proceedings, no further comment will be made.”
Barnsley boss Gerhard Struber admitted reports regarding Diaby had been a distraction for his side ahead of their Championship match against Preston on Tuesday night.
“It was distracting. The process is running but it’s not the right time to speak about Bambo,” he said after the game.
Diaby, who was born in Spain but also has Senegalese nationality, joined Barnsley last year from Belgian side Lokeren.
He has started 21 of Barnsley’s 29 Championship matches this season, scoring once.
The FA operates a research and intelligence-led anti-doping programme and it applies to all professional clubs.
The FA did not wish to comment on the matter when contacted by the PA news agency.