The injection only worked on Subjects of Ymir and had no effect on other living things. While the exact chemical composition of Titan serum is unknown, it was considered by the Marleyans to be Titan spinal fluid. It had been shown that the serums quickly evaporated after coming into contact with air; for this reason, they were always kept in tightly-sealed vials.
There were different types of serums: Rod Reiss had with him a "strongest" and an "armor" serum along with the regular serum. It is unknown if there are more types. The "strongest" serum, when ingested orally, turned Rod into a 120 m Titan, although he claimed that it makes a Titan most suited for battle (Kenny theorized that he did not follow the process correctly), and the "armor" serum gave Eren Yeager's Titan hardening abilities.
Titan serums were injected into the bloodstream, typically the inner forearm or the back of the neck; it could also be ingested. Very shortly after it is introduced into the bloodstream, the person would generate a Titan body and become a "Pure Titan." Pure Titans were mindless until they consumed a human with the Power of the Titans. Humans with the power of the Titans could take the serum again, and specialized serums can give the Nine Titans abilities they never had. After witnessing Rod Reiss turn into a disfigured 120 m Titan when he licked the "strongest" serum, Kenny Ackerman speculated that the method in which the serum enters the body matters, and if done improperly, there may be consequences. The size of the resulting Titan can also be controlled, as Sergeant Major Gross claimed that he could turn an Eldian captive into a three- or four-meter Titan, and then successfully did it.
Marley's Public Security Authorities used the injection as a life sentence against Eldians who break the law, including the Eldian Restorationists, to roam Paradis Island as Pure Titans for the rest of eternity as they eat other Eldians.
The Survey Corps have used it as a means to save the lethally wounded by turning that person into a Titan, then having it devour another with the power of the Titans to restore them to a human form.
Around 60 years before the fall of Wall Maria, Ymir and her cult of Ymir-worshipers were caught by the Public Security Authorities. They were sent to Paradis Island and were turned into Pure Titans.
In the year 832, a rebellious group of Eldians in Marley, the Eldian Restorationists, was discovered by the Public Security Authorities, thanks to Zeke Yeager. They were rounded up, sent to Paradis Island, and were turned into Pure Titans; however, one of the Marleyan officers was revealed to be Eren Kruger, "the Owl," and saved Grisha Yeager from this fate. After giving Grisha a mission to reclaim the Founding Titan, Kruger injected him with the serum so Grisha can eat him and inherit the power of the Attack Titan and reach the Walls.
On the night of the fall of Shiganshina, Grisha Yeager traveled to the Reiss Chapel to speak with Queen Frieda. After battling with Frieda in her Titan form and slaughtering most of the Reiss family, he reunited with his son, Eren Yeager, and injected him with a serum, transforming him into a Titan.
Five years later, Rod Reiss prepares to use the serum for his illegitimate daughter Historia. Upon seeing the Titan injection syringe prepared by Rod Reiss for Historia, Eren notes the identical embellished design of the syringe to the one used on himself years ago, indicating that Grisha had stolen an injection from the Reiss family. After much discussion and contemplation, Historia ends up shattering the syringe and chooses to free Eren, who was the prepared sacrifice for Historia's Pure Titan. In desperation, Rod licks the exposed serum, becoming a monstrous Pure Titan. In order to save his comrades from the collapse of the Chapel's caverns, Eren bites into an injection vial labeled "Armor," granting him the ability to harden his Titan.
At some point during the ritual for Historia's transformation, Kenny Ackerman steals another injection from Rod Reiss' bag. As he is dying, he is found by Captain Levi. Just before his death, he hands the injection over to Levi. It is later decided by the veterans of the military that Levi should be entrusted with the injection.
The injection given to Levi later becomes a subject of conflict as both Armin Arlert and Erwin Smith are brought to Levi on the brink of death. Ultimately, Levi chooses to save Armin with the injection, and Armin's Pure Titan consumes Bertolt Hoover, the Colossus Titan, and Armin inherits the power of the Titans.
In 854, the Anti-Marleyan Volunteers present a large stock of Titan serum to the Garrison, explaining they had stolen it from Marley. However, Yelena admits duplicating it will be hard since they were unable to obtain any of the materials, equipment, or experts in Titan science they need. Dot Pixis thanks the volunteers, while he begrudgingly arrests them over uncertainties regarding their leader.
- While only the "Armor" vial read by Eren has been identified officially in the story, the labels of the three Titan injection vials that have been introduced each bear legible katakana:
- "Strongest Titan" (サイキョウノキョジン Saikyō no Kyojin?) - The one Rod chose for Historia and which he ended up ingesting.
- "Armor Braun" (ヨロイ ブラウン Yoroi Buraun?) - The one Eren ingested and which granted him hardening abilities.
- "Normal Titan" (フツウノキョジン Futsū no Kyojin?) - The one Kenny gave to Levi and which was injected into Armin.