Here are some things to know about this list going in:
A) I used precise, secret, and forbidden mathematical algorithms combining difficulty, lore, move set, and boss design.
B) YES! This does include the DLC bosses
C) I did not include Demon Souls or Bloodborne in this list, because they don't count. And if you think they do, you are dead wrong. Also if this post does well I need an excuse to write up another one.
D) When making lists like these, some folks like to prattle on and on about how hard it was to make it because of how wonderful each of the entries are. Well, for me, it was idiotically easy. Every single boss on this list is clearly greater than the last, no ifs, ands, or big stinky butts about it. You animals.
40. The Lost Sinner
You'd better not be a horny teenager, because this boss looks like she's been ripped straight out of a slasher flick. This is especially true if you haven't illuminated the room by setting the oil tracks outside of her chamber ablaze which are unlocked by obtaining the Bastille Key from the Belfry Gargoyles (have no idea what I'm talking about? TURN AWAY NOW). She is one of the more memorable bosses in DS2 (which has a reputation for having too many lousy adversaries). Her move set is almost balletic with swift sweeping attacks and furious stabs that can close the gap between you and her even if she's on the other side of the room.
39. The Gaping Dragon
If this list were based solely on boss design alone, the Gaping Dragon would be in the top 5. If this were a list of the top screaming toothed genital monsters of all time, it would be number one. This battle can last a while due to the dragon's high defense leading to a long and exhausting fight. A general rule of thumb in these games is that if it's a massive monster like this one, get behind it and get ready to slice some butthole. Be careful, it can Space Jam up into the air and squash you like a goomba in a bounce house full of 9 year olds.
38. Seath the Scaleless
Well whaddya know, it's another dragon! Specifically, he's the dragon that betrayed the other dragons and then went insane via his own research in his archives. As you can see, the lore on this boss is super cool, and so is his appearance. The fight itself is... not as mind blowing. Once you shatter the crystal that keeps him immortal, you just get up in his nuts and whack away. Yes, that's exactly how I intended that to sound. He can also pulse out crystal shards on the floor around you which can curse you. Curse is like the AIDS of Dark Souls, people ignore it until it's too late and you have to make a Tom Hanks movie. Instant death and you only have half your maximum health going forward unless you pay a buttload of souls for a purging stone.
Side note: the boss run to get to him is an absolute pain. Screw you, Crystal Cave, and your giant clam monsters too!
37. Looking Glass Knight
Sporting a fancy mirror shield and a creepy doll face, this warrior hits hard with a lightning infused sword that really makes you want to keep your distance. What makes this classy doctor stand out like a Snickers bar in a swimming pool is that he can summon NPCs or even REAL ON-LINE PLAYERS into the fight. If you don't play these games (in which case you may want to consider signing up to be a professional baby), then let me tell you, real life players can be devastating to fight if they have a really high-level build. Can you imagine taking on this bad boy when all of a sudden a dude named Noobkiller420 hops out at you dual wielding red rust swords? It feels like having to win an MMA tournament while there's a coyote actively biting your dick.
36. Dragonslayer Armour
When you first enter the arena, the sight is breathtaking. Looming over the Dragonslayer floats a massive flying creature that looks like it's made out of veins. These creatures are called the Pilgrim Butterflies and they are the puppeteers of the Dragonslayer who is actually an empty set of armor. With epic music blaring in the background, the boss slams its massive shield and axe down on your whole face relentlessly. The second phase gets even more nuts when the butterflies start vomiting huge blood balls at you alongside the Dragonslayer's melee. Side note: his dragonslayer greataxe is a phenomenal weapon that's so intoxicatingly powerful that it makes you feel like you're perpetuating a cycle of abuse, but like, in a good way.
35. The Rotten
The Rotten is one of the coolest boss designs in Dark Souls 2 because it's made out of people. PEOPLEEEEEEE! That's right, I made a Soylent Green reference. What of it?! You can find this hunk of chunks within the Black Gulch past a bunch of big gross worms. Its attacks mirror those of a Dark Souls boss named Gravelord Nito which we will get to later in this list, and rumor has it that he is a reincarnation of said boss. Another cool detail is that there is a lone undead on The Rotten's shoulder that imitates its movements which would suggest that the little guy is controlling the big lug. I like to name that guy Larry, and have made it my duty to edit all the various Souls wikis until that name becomes an undisputed part of the lore.
34. Asylum Demon
Welcome to your tutorial. One of the most memorable moments in the series takes place right at the top of Dark Souls when you run into a room wielding nothing but a broken sword. After having barely done anything in the game, all of a sudden you're in a boss fight with this gnarly ogre. It's one of the most important lessons in the game as you quickly learn that there's no shame in high tailing it out of there and come back when you're better equipped. This teaches an essential fundamental of the series: just run away, man, it's not worth it. Also, this guy has a butt stomp that would put Niki Minaj to shame.
33. Sif, the Great Grey Wolf
It's a wolf with a giant sword in its mouth. What more do you want from me? Maybe some super sad back story stuff? You got it! Sif was Sir Artorias' companion who now guards his grave since the knight's passing. In the DLC content, you come to find out that Artorias succumbed to the abyss long ago, but before he did so, he created a protective barrier around Sif to save him from corruption. If you go through the DLC content before you fight Sif, an extra cut scene happens in which the wolf will recognize your scent and then howl at the moon in despair. To make matters worse, when you get the wolf low enough in health, the poor little doggie starts limping around. Could you at least have made this one optional, Miyazaki?
32. Gravelord Nito
They should rename this guy Bones For Days. I keep writing From Software to make this happen but THEY NEVER GET BACK TO ME! Nito looks like the most ripping of metal album covers. He's referred to as "the first of the dead" and can mess you up with his gravelord sword. Not only that, but he is surrounded by a bunch of his skeleton buddies who are also attacking you, all looking pleased as punch while they do it too. Did somebody say giant skeletons? Of course not. Who would say a thing like that? Regardless, giant skeletons will also attack you during this fight. The only thing that can save you is Nito himself whose attacks hurt both you and his minions. It's a big bone party in the Tomb of the Giants tonight, so come on down!
31. Smelter Demon
This asshole. I know people who quit the game after their hundredth run at the Smelter Demon. A little ways into the fight, he seppukus himself to turn his big awful sword into a big awful flaming sword. With every hit, he becomes faster and more powerful. Everybody hates the Smelter (coming to CBS this fall). I personally had to summon in help to fight this bag of flaming dog doo and even that didn't work. Luckily the boss is optional so it's recommended that you go level up a whole bunch and come back later. Or maybe just don't come back at all and go on to live a happy, peaceful life.