An internet watchdog based in Canada, Citizen Lab, said the surveillance software is only sold to governments.
Last week, Citizen Lab also said that 12 journalists and human rights lawyers had been targeted.
At the time, the Mexican president, Enrique PeÃ±a Nieto, described accusations that the government was involved as false.
Mr PeÃ±a Nieto said surveillance software was only used for matters of national security and for fighting organised crime.
He added that he had ordered an investigation by the attorney general's office.
Citizen Lab said the head of the Conservative National Action Party (PAN), Ricardo Anaya, a PAN Senator, Roberto Gil Zuarth, and the party's communications secretary, Fernando Rodriguez, were targeted.
"There is strong circumstantial evidence implicating the government of Mexico", the watchdog said.
The software, known as Pegasus, was sold to Mexican federal agencies by the Israeli company NSO Group on the condition that it only be used to investigate criminals and terrorists.
It is usually sent in a text message to a smartphone.
If the person taps on it, the spyware is installed, and huge amounts of private data - text messages, photos, emails, location data, even what is being picked up by the device's microphone and camera - are hacked.
Three senior opposition politicians in Mexico have been targeted by spyware on their mobile phones, researchers say.