= Rule of Thumb = {F5928403, size=full} = Example = {F5928405, size=full} = Limit = (IMPORTANT) If you're object size is below the limit of diffraction (see [[ https://en.wikipedia.org/wiki/Diffraction-limited_system | Abbe diffraction limit ]]) , ie roughly 250 nm, no need to acquire images with pixel size of 2 Å ! (NOTE) Please use this [[ https://svi.nl/NyquistCalculator | Nyquist calculator ]] to get the smallest size of pixel which is meaningful with your microscope capabilities. You can lower the obtain value by 1.5 and still get nice imaging. = Object size and Intensity measurement = (IMPORTANT) With an **inadequate pixel size**, measuring **size** and **intensity** of your object will be **bias**. In the example below, you can see that a translation of only **half-pixel** would make the object twice larger and twice dimmer the Object in respect to pixel in one case and without consequence in the other one. {F5929467, size=full} = Range = {F5928589, size=full}

(NOTE) Do not forget, it's [[https://c4science.ch/w/bioimaging_and_optics_platform_biop/teaching/probes/ | A Matter of Scale]] = Rule of Thumb = {F5928403, size=full} = Example = {F5928405, size=full} = Limit = (IMPORTANT) If your object size is below the limit of diffraction (see [[ https://en.wikipedia.org/wiki/Diffraction-limited_system | Abbe diffraction limit ]]) , i.e. roughly 250 nm, there is no need to acquire images with pixel size of 2 Å ! (NOTE) Please use this [[ https://svi.nl/NyquistCalculator | Nyquist calculator ]] to obtain the smallest meaningful pixel (voxel) size that matches your microscope's capabilities. If needed, you can lower the obtained value by 1.5 (in XY) and still obtain high quality images. = Object size and intensity measurement = (IMPORTANT) With an **inadequate pixel size**, the measured **size** and **intensity** of your object will be **bias**. In the example below, you can see that a translation of only **half a pixel** would make the object twice as large and twice as dim in the case of an inadequate pixel size (left), and be without consequence with optimal sampling (right) {F5929467, size=full} = Range = {F5928589, size=full} (IMPORTANT) What is true for XY, is also true for Z. Please do not make z-stack acquisition, with a pixel size of 40nm and a z-step of 2um!

(NOTE) Do not forget, it's [[https://c4science.ch/w/bioimaging_and_optics_platform_biop/teaching/probes/ | A Matter of Scale]] = Rule of Thumb = {F5928403, size=full} = Example = {F5928405, size=full} = Limit = (IMPORTANT) If you'rer object size is below the limit of diffraction (see [[ https://en.wikipedia.org/wiki/Diffraction-limited_system | Abbe diffraction limit ]]) , i.e. ie roughly 250 nm, there is no need to acquire images with pixel size of 2 Å ! (NOTE) Please use this [[ https://svi.nl/NyquistCalculator | Nyquist calculator ]] to getobtain the smallest size ofmeaningful pixel which is meaningful with(voxel) size that matches your microscope's capabilities. YIf needed, you can lower the obtained value by 1.5 (in XY) and still get niceobtain high quality imaginges. = Object size and Iintensity measurement = (IMPORTANT) With an **inadequate pixel size**, the measuringed **size** and **intensity** of your object will be **bias**. In the example below, you can see that a translation of only **half- a pixel** would make the object twice larger and twice dimmer the Object in respect to pixel in one case and without consequence in the other one.as large and twice as dim in the case of an inadequate pixel size (left), and be without consequence with optimal sampling (right) {F5929467, size=full} = Range = {F5928589, size=full} (IMPORTANT) What is true for XY, is also true for Z. Please do not make z-stack acquisition, with a pixel size of 40nm and a z-step of 2um!