1. Name: Pitch Black
Specialty / Start with: You can not see his heart; Rooms are dark; No stores
Objective: Mom
Available: immediately
2. Name: High Brow
Specialty / Start with: Number One, E. coli, Butt Bombs, Flush! and Petrified Poop
Objective: Mom
Available: immediately
3. Name: Head Trauma
Specialty / Start with: Tiny Planet, Soy Milk, Small rock and Iron
Bar
Objective: Mom
Available: immediately
4. Name: Darkness Falls
Specialty / Start with: Pentagram, Dead Bird, Whore Of Babylon, Razor Blade, Sacrificial Dagger and Dark Matter
Objective: Satan
Available: enable "It Lives"
5. Name: The Tank
Specialty / Start with: Lard, Infamy, Yum Heart and Thunder
Thighs
Objective: Mom
Unlock "Magdalene": Available
6. Name: Solar System
Specialty / Start with: Transcendence, Distant Admiration, Forever Alone, Halo of Flies. You start out as "Lord of the Flies". Damage can be caused only by flying.
Objective: Mom's heart
Unlock "Transcendence": Available
7. Name: Suicide King
Specialty / Start with: Ipecac, Mr. Mega, My Reflection
Objective: Isaac
Available: enable "It Lives"
8. Name: Cat Got Your Tongue
Specialty / Start with: Guppy's Head, Tail Guppy's and Guppy's
Hairball.
You start as a guppy. Damage can be caused only by blue flying and his hairball.
Objective: Mom
Available: yourself transform into a run in "Guppy".
9. Name: Demo Man
Specialty / Start with: Dr. Fetus, remote detonator, Matchstick
Objective: Mom's heart
Unlock "Dr. Fetus": Available
10. Name: Cursed!
Specialty / Start with: Yum Heart, Liver Raw, The Compass, Treasure Map, Map Blue, Child's Heart. Make entering and leaving spaces harm.
Objective: Mom
Unlock "Magdalene": Available
11. Name: Glass Cannon
Specialty / Start with: Epic fetus, Loki's Horns
Objective: Satan
Available: "Epic fetus" (Challenge # 19) and "Loki's Horns"
Unlock
12. Name: When Life Gives You Lemons
Specialty / Start with: Lemon Mishap, 9 Volt, Habit and 1 Lemon
Party pill
Objective: Mom
Available: immediately
13. Name: Beans!
Specialty / Start with: Pyro, The Bean, The Black Bean, 9 Volt, Butt Bombs. You can not shoot tears. Damage you do only by bombs and "The Bean"
Objective: Mom
Available: immediately
14. Name: It's in the Cards
Specialty / Start with: Deck of cards, starter deck, The Battery, 9 volt and 2 random tarot cards.
Objective: Mom
Available: immediately
15. Name: Slow Roll
Specialty / Start with: Polyphemus, My Reflection, Cupid's Arrow
Objective: Mom
Available: immediately
16. Name: Computer Savvy
Specialty / Start with: Technology, Technology 2 and Spoon
Bender
Objective: Mom
Available: immediately
17. Name: Waka Waka
Specialty / Start with: Strange Attractor, Anti-Gravity
Objective: Mom
Available: immediately
18. Name: The Host
Specialty / Start with: Spider Baby, The Mulligan and The Tick
Objective: Mom
Available: immediately
19. Name: The Family Man
Specialty / Start with: Brother Bobby, Sister Maggy, Dad's Key, BFFS! and Rotten baby. You can not shoot tears. Damage only make the bombs and the companion of Isaac.
Objective: Isaac
Available: "Dad's Key" and activate "It Lives"
20. Name: Purist
Specialty / Start with: no items to start
Objective: Mom's heart
Available: defeat "Mom's heart" for the first time