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After your individual exposures, known as subframes (subs) have been captured, they then go through a process known as calibration. This is neccesary to remove any artifacts added to the image by your imaging equipment. The most common artifacts are hot pixels, uneven illumination / vignetting and dust shadows.

Calibration is done by taking sets of exposures known as calibration frames. These are then subtracted from your images, thus removing any artifacts. These artifacts are regarded as noise, and most of them can be regarded as fixed pattern noise (FPN), as the noise will be the same in all frames for a particular imaging session, provided the equipment is not altered during the session.

There are 4 types of calibration frame. These are dark frames (Darks), bias frames (Bias), flat frames (Flats), and flat dark frames (Flat Darks) With the exception of flat darks and bias (flat darks are only applied to your flats, and bias frames are only applied to your lights if you don't apply dark frames.), these are applied to the individual images that you have captured, before any stacking or alignment is done.

Lights are your images....the shots the camera takes.

Bias frames are a record of your system's inherent noise with zero exposure.
They are of the minimum exposure length you can get, if possible, zero.
They are taken with no light entering the scope.

Darks are a record of the noise when exposing and are very temperature dependent. The higher the temperature, the more noise you get, in the form of hot pixels.
They need to be of the same length and taken at the same temperature as the lights.
They are taken with no light entering the scope. This is M57 with no darks applied, and M57 with darks. (The black marks are caused by dust near the sensor.) Dark frames also contain bias information, so bias frames don't need to be subtracted from the lights if darks are being subtracted.

Flats are a record of all the optical irregularities in the system, the most common being vignetting, where the image is lighter in the centre than the outside, and dust particles (Dust particles create small donuts when they're near the sensor, and larger ones the further away they are). Flats also reduce fixed pattern noise in the form of uneven sensitivity of the pixels making up the ccd detector. Both dust particles and vignetting can also be regarded as a type of FPN, albeit of a temporary nature as they will change when the optical trian is altered.

Very Important!!.......Flats must be taken with the optical train exactly as it was when the lights were taken...the camera must absolutely not be moved. It is generally agreed that focus shouldn't be changed either, but in the real world this can be a bit tricky, as focus tends to shift over the course of the night as the temperature changes, and it's not practical to shoot a new set of flats every time you adjust focus....you'd have no time to image! In practice, most of the issues that are solved by flats are not too focus critical. These are dust particles/smears etc, and differences in pixel sensitivity. Dust particles on focal reducers, filters, and optical windows in front of the chip do not change distance with focus, as your focal reducer, filter wheel and camera all move as one unit. What is affected by focus changes is vignetting, but not generally by much, and any dust on the objective lens is so far out of focus as to be invisible. Although it's probably best practice to shoot flats on the night you are imaging, I often do mine the following day, as at 6am, after a 12 hour overnight session in the depths of winter, I'm not generally in the mood to shoot flats! As my optical train remains unaltered for weeks at a time, I find that once I have a good set of flats, I can use them for anything I take for the next few sessions, although eventually dust can creep in and make a new set of flats neccesary.

This is an example of what can happen if the optical train has been moved when the flats are taken, and is an absolute pig to fix!! (The CCD in the camera was adjustable, and wasn't locked down, so it had moved between the imaging session and when the flats were taken!)

A separate set of flats will need to be made for each filter, as the dust particles will be different for each of them. They are taken using an evenly illuminated lightsource, commonly a lightbox, or electroluminescent (EL) panel.

Flats need to be bias subtracted in order for the correct subtraction calculation to be made. If they aren't, they will either over, or under compensate.

There are also Flat darks. These are dark frames of the same exposure length as the flats and, if used, are applied to the flats.

I find that the flat exposures are normally quick enough not to worry about noise buildup, and bias frames handle the rest, so I don't do flat darks.
I apply bias subtracted flats and temperature matched darks to my individual subs. The master bias, dark and flat are comprised of more than 20 individual sub exposures. If you don't do enough calibration frames you will end up adding noise to your images.
I combine my calibration frames into master darks, flats and bias using an 'average' combine algorithm and do not use any alignment routine.