Hi!
If you need help with finding all characters,or you don't know if you got them all, this walkthrough may be for you! This will (hopefully) help you with all the characters. Everything will be listed in the order of their number. Both Japanese and English versions of the game have the same locations, so this guide works for both versions.

Before I start however, if you wish to find out whether you have everyone unlocked, go to your Omega Factor (by pressing SELECT in while in a level) and look for Sharaku (He is located in a big icon to the right; he is marked as "EVIL"). If he is colored, you have all characters.

Before we start, quick explanations. We will ignore the "?" and "!" that appears in the Omega Factor (Dr. O'Shay explains it to you in the beginning of the game). There are 4 versions of people. Normal, the one you see the most, big, the big ones, hidden, the ones that are hidden, and then the middle sized, which marks the end of half the game. Specifically, I wish to talk about the big types. The big types will start out as Grey. This will not let you upgrade your powers. So for the big ones, they will appear twice. The way to unlock the grey version will be written as (Grey), while the colored versions will be written as (Colored). Note that you can not color any characters in the Birth section. You must have Stage Select unlocked.

There are only 2 middle sized icons; Phoenix & Garon (46 & 43). Having Phoenix means you finished the first half. Having Garon means having finished last half. Thought I'd clarify this. Anyway...


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Sections;
00-Astro
01-Dr. O'Shay
02-Wato Chiyoko
03-Detective Tawashi
04-Amazing Three
05-Big X
06-Rainbow Parakeet
07-Wally Kisagari
08-Skunk Kusai
09-Kinsankaku
10-Mr. Tokugawa
11-Atlas
12-Rag
13-Duke Red
14-Nuka
15-Black Looks
16-Deadcross
17-Rock
18-Sapphire
19-North
20-Denkou
21-Brontus
22-Epsilon
23-Pluto
24-Zoran
25-Blue Knight
26-Alejo
27-Abercrombie
28-Kennedy
29-Boon
30-Magma
31-Daichi
32-Prime Rose
33-Drake
34-Jetter Mars
35-Fumoon
36-Hecate
37-Unico
38-Don Dracula
39-Ham Egg
40-Pinoko
41-Black Jack
42-Pook
43-Garon
44-Sharaku
45-Dr. Tenma
46-Phoenix


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  00- Astro
If you do not have him unlocked, please consider sending a screenshot to me as proof.

  01-Dr. O'Shay
When you start a new game, you will be in a tutorial level. Simply complete this level.
  If you do not have Dr. O'Shay unlocked, you must finish the Birth section of the game and unlock Stage Select. From here on just go to Stage 0-1 and finish the stage. You will meet Dr. O'Shay and unlock him in your Omega Factor.


  02-Wato Chiyoko
Simply beat Stage 1-4.


  03-Detective Tawashi
Defeat North in Rebirth Stage 7-1.


  04-Amazing Three
In Stage 2-3, defeat the first boss you will find.


  05-Big X
After defeating The Amazing Three in Stage 2-3, once you enter the cave, just head to the right. You will find a character who will fight you. Defeat him and you have unlocked Big X.


  06-Rainbow Parakeet
After defeating The Amazing Three in Stage 2-3, head into the cave and enter the first hole you will find. From there, head left. You will find him.


  07-Wally Kisagari
In Stage 2-3, enter the first hole, head right, and once you can't head right anymore, head left, up, left and you will find Wally. Talk to him.


  08-Skunk Kusai
Defeat Denkou in Rebirth Stage 7-2.


  09-Kinsankaku
Defeat Brontus in Rebirth Stage 7-3.


  10-Mr. Tokugawa
Defeat Epsilon in Rebirth Stage 7-4.


  11-Atlas
Defeat him in Rebirth Stage 3-5.


  12-Rag
Simply beat Stage 3-5.


  13-Duke Red (Grey)
Simply beat Stage 3-5.


  13-Duke Red (Colored)
First, you must defeat Pluto in Rebirth Stage 7-5. Then, head to Stage 4-1. Finish it. Abercrombie will tell you where the hide-out of Duke Red is. Now, enter Stage 4-2, and stick to the very bottom of the level. You will enter a building once you reach it. You must now fight Blue Knight. Defeat him and you will unlock Duke Red.


  14-Nuka (Grey)
Simply beat Stage 4-1.


  14-Nuka (Colored)
Simply beat Rebirth Stage 2-6.


  15-Black Looks
Simply beat Stage 5-1.


  16-Deadcross
Simply beat Stage 5-4.


  17-Rock
Simply beat Stage 6-4.


  18-Sapphire
Defeat Sharaku in Rebirth Stage 6-4.


  19-North
Defeat him in Stage 7-1.


  20-Denkou
Defeat him in Stage 7-2.


  21-Brontus
Defeat him in Stage 7-3.


  22-Epsilon
Defeat him in Stage 7-4.
  Fun fact: despite the fact he has breasts, he is considered male in-game. Strange design for a male, I know.


  23-Pluto
Defeat him in Stage 7-5.


  24-Zoran
Defeat Pluto in Rebirth Stage 7-5.


  25-Blue Knight
First, you must defeat Pluto in Rebirth Stage 7-5. Then, head to Stage 4-1. Finish it. Abercrombie will tell you where the hide-out of Duke Red is. Now, enter Stage 4-2, and stick to the very bottom of the level. You will enter a building once you reach it. You must now fight Blue Knight. Defeat him and you unlock Blue Knight.


  26-Alejo
In Stage 1-2, once you have defeated the stop/go robot in the beginning of the stage, destroy the top-right window. He will fall and you can talk to him. You have unlocked him.


  27-Abercrombie
Simply beat Rebirth Stage 4-1.


  28-Kennedy
Immediately upon entering Stage 5-2, head left and destroy the trashcan that you will find.


  29-Boon (Grey)
After finishing Rebirth Stage 1-4, from the Stage Select screen, enter Stage 0-4. You will unlock Boon.


  29-Boon (Colored)
This requires you to do multiple things. This is also required at some point of the story to advance. First, you must unlock Magma. (See number 30) After that, once you have Stage Select (You can still get Magma if you already have Stage Select), talk to Boon and unlock him in Grey. (Finish Stage 1-4, enter Stage 0-4)
Once you have Boon in Grey, head to Stage 6-3. Go to the end and blow up the second door on the train. You will find Don Dracula, and he will tell you something about Sharaku's soul. Now head back to Stage 0-4. You will unlock Boon.


  30-Magma
In Stage 2-5, use the background as a clue. Once you can move, look at the background. See these dark spots? Wait until the third dark spot. Punch in the topmost area of the dark area. You will find Magma, the robot by the one and only Kami aka Dende (who does not exist in this game) aka God.


  31-Daichi
After obtaining Pinoko (see number 40) and defeating the Roboid in Stage 0-3, you will automatically head to Stage 3-5. Defeat Atlas while keeping Prime Rose safe and you will unlock Daichi.


  32-Prime Rose
After defeating the Roboid in Stage 0-3 and defeating Atlas one last time, you will unlock her.


  33-Drake (Grey)
In Rebirth Stage 1-1, finish the first wave and head right.


  33-Drake (Colored)
After defeating the Roboid in Stage 0-3 and defeating Atlas one last time, you will unlock him.


  34-Jetter Mars
After the first Enemy Wave in Stage 3-1, head right and jump up. You will notice how there are so many blocks. You can destroy the second-to-last block at the top that you will find. Destroy it. You will now find Jetter Mars and unlock him.


  35-Fumoon
In Stage 3-3, head right in the beginning until you can't head right. Before heading up & left, Jump up. You will find this random floating platform. If you punch at the very left on the platform, you will notice Astro is actually punching something. After punching it several times, you will find an alien, known as Fumoon.


  36-Hecate
In Stage 3-4, after defeating the 8 enemy waves, you can just head right to finish the level and head to Stage 3-5. Before you do, however, head up and then left. You will find Hecate.


  37-Unico
After defeating the first wave of enemies in Stage 6-2, fly up, up, up and up. You will find something. Go in it and destroy the statue.


  38-Don Dracula
At the end of Stage 6-3, you will find the train from Stage 5-4. Simply blow up the second door, and boom, Don Dracula appears.


  39-Ham Egg
After unlocking Pinoko (see number 40), head to Stage 0-3 and defeat the Roboid. Note that after obtaining Ham Egg, you can not use Stage Select until you finish the game. You are nearing the very end of the game.


  40-Pinoko
After unlocking Black Jack in Grey (see number 41), head to Stage 2-3 and meet up with Rainbow Parakeet. (see number 06) After you have talked to him, head to Stage 2-6. Instead of fighting Pook, a cutscene will play and you will unlock Pinoko.


  41-Black Jack (Grey)
After you have unlocked Duke Red in colored, (see number 13 (25 also works)), go to Stage 1-2 and unlock Alejo. Even if you already have him. (See number 26) After that, head to Stage 1-1, and meet with Drake. Once you've talked to him, you must head to Stage 3-5 and defeat Atlas. Now head to Stage 5-2 and talk to Kennedy (See number 28). You will now unlock a new Stage. Head for Stage 0-3.
NOTE: You must have upgraded your Sensors to Level 3!!! Or else you will not unlock Black Jack! Look for characters that you could still unlock and upgrade Sensors to Level 3 if you haven't gotten that Level. Once you have the required Sensor level, head back to Stage 0-3.


  41-Black Jack (Colored)
After unlocking Pinoko (see number 40), head to Stage 0-3 and defeat the Roboid.


  42-Pook
Simply defeat him in Stage 2-6.


  43-Garon
After beating the Birth section, play through Rebirth Stage 1-1 to 7-5. Then head to 4-1. Finish it. Head to 4-2. Stay at the bottom. You'll find Duke Red. Defeat Blue Knight now. Head to 1-2, meet with Alejo (see number 26), now go to 1-1 and meet with Drake. Make sure you have Boon colored (see number 29).
Go to Stage 3-5. Defeat Atlas and go to 5-2 to meet with Kennedy. Head to Stage 0-3 with Sensors level 3, and then head to 2-3 and meet with Rainbow Parakeet (see  number 06). Now go to Stage 2-6, then 0-3, defeat Roboid, then defeat Atlas, and finish the next few stages. Now defeat Garon in Stage 8-4.


  44-Sharaku (Grey)
Defeat him in Stage 6-4.


  44-Sharaku (Colored)
Obtain every person in your Omega Factor. After this, head to Stage 8-5 and finish the stage. After the credits you will see a cutscene. After that cutscene, a "THE END" screen will appear. Now simply wait a minute or two. After having the patience, a cutscene will play, the story truly ends in a nice way, and you obtain Sharaku, fully colored! Congratulations, you have completed the game 100%!


  45-Dr. Tenma
Once you have unlocked Stage Select (AKA Rebirth), visit Stage 0-1. You may have already done the tutorial, but remember, Astro is smarter this time. When you finish the stage and talk to Dr. O'Shay, Astro will ask about his father. You will now unlock Stage 0-5. Visit Stage 0-5.
After visiting the Stage, head to Stage 6-2, finish the first wave of enemies, fly up and destroy the statue inside the ...whatever that is. You will find Unico, and they will grant you a wish. Now head back to Stage 0-5. Astro will remember more, but Dr. Tenma is there as well, and he will explain to you why Astro was created and his story when Astro was there. And how Tobio was killed. You will now unlock Dr. Tenma, Astro's/Tobio's father.


  46-Phoenix
Defeat Pluto in Stage 7-5.



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Edit as of 12/09/2016:
Fixed capitalization. Oops.

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Edit as of 23/09/2018 (yeah, two years later, ikr who edits posts so late? xd):
Fixed some if not most grammar issues, clarified some things. Should be slightly easier to follow!
Also fixed a spelling mistake in the 12/09/2016 edit log. "Capitilization"? Really? God I used to be so bad at everything. Oh how I've grown since then. I love you guys <3