
Continuity mistake: When Norman is getting ready to test himself, he lies down on the bed, fastens himself in and the doctor goes to the computer. However, when it shows him being brought in to the chamber he has several electrodes connected to his chest and head. (00:15:20)
Suggested correction: At first the electrodes are beside Norman's head. While the doctor is fixing him in place, at one point camera focuses on his face. Later when he's stepping back for the PC, the electrodes can actually be seen stuck on his body. You can see them if you have an HD copy. So the camera focusing on his face is indicating that the doctor's doing whatever necessary for the process.

Continuity mistake: When Peter shoots his web at his bedroom lamp and pulls it across the room, it smashes against the wall and breaks. But when Aunt May is talking to Peter from the door seconds later, the lamp is back on the dresser in one piece. (00:32:15)

Continuity mistake: When Mary Jane is being mugged by four men, Spider-Man throws two of the men into two windows behind Mary Jane. The shot switches to Spider-Man beating up the other two guys. When it cuts back to Mary Jane the two windows are intact. (01:17:00)
Factual error: In the museum, Peter and his friends are real-time videos of live, moving biological specimens shot through 'the largest electron microscope on the Eastern seaboard'. You can't film live specimens in an electron microscope. The electron beam only works in a vacuum chamber, in which the specimen - invariably dead - is held. The microscope is identified as a scanning electron microscope - current technology - and nobody can fire an electron stream through air. The electrons will collide with gas molecules and scatter, ruining the image.
Revealing mistake: When Peter Parker first discovers he can climb up walls (while in citizen clothing), he tests it out by climbing up a building in an alley. During this scene, you can clearly see his clothes hanging out forwards, indicating that the scene was filmed with him crawling along a floor horizontally. (00:24:50)
Other mistake: The scene at Columbia University was filmed on an unseasonably warm spring day, however, the costume department had provided the high school extras with cold-weather clothing. The real Columbia University students can be seen in the background wearing shorts and t-shirts by contrast. (00:05:35)
Visible crew/equipment: When Spider-Man is rescuing Mary Jane from falling to her death from the balcony, he catches her and spins his web to the balcony to save her and they bungie to just inches from the ground. As they are preparing to bounce back up the shot rotates 90 degrees and you can see the counter weight that pulls them back up coming down to the ground. It's a weird one, this - there's a sound effect as the weight comes into view, so it's meant to be there (or else the sound's just a cover), but it can't be anything he attached his web to, as it was stuck straight onto the balcony. If it's falling, they should be too - his web's not pivoting around anything. (01:06:50)
Suggested correction: They bounce back because of the elasticity of the web. That falling block is just a falling piece similar to the long one that is already on the ground, it's not a counter weight.
Suggested correction: I genuinely don't understand this mistake. It's just a piece of debris falling. It looks absolutely nothing like a "counterweight," nor is it big enough to be. It's the exact same color, texture and relative size as the larger pieces of debris on the ground below them. (Furthermore, you can see other, smaller pieces of debris falling around them during the two shots as well.) And the reason they spring back up is because the web is somewhat elastic in quality. This is shown multiple times throughout the movies. He stuck the web to the balcony, and they sprang back up as bits of debris continued to fall. There's no mistake here.

Continuity mistake: In the final battle, the blood on Spider-Man's chin and under his nose keeps disappearing and reappearing. (01:42:15)
Other mistake: When Spider-Man saves the baby from the burning building, it's amazing that the baby and the blanket are so clean - no soot, smoke damage, charring, etc. The firefighters have obvious soot around their noses from trying to breathe in that area. Did Spidey manage to give the baby a bath before presenting it to momma? (01:19:40)

Continuity mistake: When Jameson is flipping the pictures of Peter's first batch of photographs, you can see that the 3rd picture he looks at changes into an entirely new photograph, when he puts it down. The one he's looking at is Spider-Man on the side of a building, but when it's on the desk it's Spider-Man kicking a thug in the face. The one he then looks at changes too - it was about to have a bit of sky on the left, but then turns into a photo of a street from above, with no sky visible. (00:58:45)

Continuity mistake: When Peter is talking with Mary Jane outside the diner where she works, watch the positioning of the shadows in the background. As the camera shoots from Peter's point-of-view, the sun is casting shadows up the street. When the shots change to MJ's point-of-view, the shadows are pointing down the street, as if the sun had just moved ahead several hours. (00:57:30)
Other mistake: When the tram with all the kids is falling, the kids are yelling. But the adult behind them isn't yelling and looks rather bored. (01:38:10)

Continuity mistake: When Spider-Man is chasing the crook with the gun, the crook shoots 7 times through the roof of the car. Shortly afterwards you can see 9 bullet holes in an overhead shot. As the scene moves along there is another overhead shot (just after Spider-Man jumps the bridge) - no bullet holes. Seconds later the bullet holes return. (00:44:40)
Factual error: When Harry is talking to Mary Jane on the phone, she hangs up on him and his cell phone produces a dial tone. Cell phones do not have a dial tone. (01:08:25)

Continuity mistake: When Peter Parker confronts Uncle Ben's killer in the warehouse, after Parker slams him through the glass windows in the door, the killer drops his gun and pulls out a knife. After he is disarmed of the knife, Peter kicks him up against the wall. At this point the killer has the gun back in his hand. (00:47:50)
Continuity mistake: When Peter first learns he can climb walls after running out of school, his shoes are different when he is on the wall compared to when he is on the roof. (00:26:20)

Continuity mistake: When Peter is drawing up ideas for his costume the hand is that of comic artist Phil Jimenez, current artist of Wonder. Phil Jimenez is right handed and Tobey is left handed. In one of the cuts the pen is in Tobey's left hand but it shows him drawing with his right. (00:31:25)
Continuity mistake: During the kiss scene the camera changes view back and forth. From one angle it's raining, in other it is not, then at the end it's pouring as if in a storm. They had to do that scene at least 3 times. (01:17:50)

Continuity mistake: In the cafeteria scene, Mary Jane slips on some spilled orange juice that causes Peter to catch her. A short while later when Peter gets up to leave the cafeteria, running away from her enraged boyfriend, the spilled orange juice is not there. (00:22:15)

Continuity mistake: In the final cemetery sequence, Peter and Mary Jane square off for a little heart to heart, with her touching his face tenderly with her black leather gloves. The camera intercuts between front views of both: in hers, her fingers are touching his ear lobe, in his, they are a inch below his ear lobe. In one quick cut of hers, the hand has disappeared completely, then in mid-sentence as they cut back to Peter, it's there again. (01:49:20)