8 Note about electron dose

Since we know the conversion rate of electron to counts and pixel size for each magnifications, we could use these two parameter to figure out the relationship between the mean counts on CCD and dose on specimen. It is as following:

Mean_Counts = Dose * Conversion_Rate * Pixel_Size * Pixel_Size

For example, if you take images at mag of 19500X(ScreenDown), the pixel size at specimen level is then 0.488nm. If you want 20e/A2 as target dose, you can then adjust your beam condition and exposure time to get your CCD image with Mean Counts of

Mean_Counts = 20 * 24 * 4.88 * 4.88 = 11431

It is important to point out that in default setup for SerialEM the readout count on your screen is only half of the original counts. This is done by checking "divided by 2 for 16 bits camera" under Camera menu. It is believed that this will remove noise more effectively, in the case of dark image background drifts sometimes. Quoted here is from SerialEM help file "Data can be divided by 2 as soon as they are acquired, by selecting Divide 16-bit By 2 in the Camera menu. They will thus never appear to be unsigned integers, making the options for treatment of 16-bit data irrelevant. If the camera noise level is high enough to make this an acceptable procedure, then it has the additional advantage that it will preserve negative values in gain-normalized images. Such values can occur and be meaningful in images with very low counts if the bias of the camera drifts downwards after taking a dark reference.". Therefore, if you measure your count in serialEM for dose purpose, you either uncheck "divided by 2 for 16 bits camera" and read the counts directly or otherwise use half of the counts in above equations.

To make it clear, if the menu "divided by 2 for 16 bits camera" is being checked in SerialEM, one should use the following equations:

Mean_Counts = 0.5 * Dose * Conversion_Rate * Pixel_Size * Pixel_Size 
Mean_Counts = 0.5 * 20 * 24 * 4.88 * 4.88 = 5715