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Choosing a Microscope Camera


diff erent areas of the specimen. A good way to think about this is if my lowest detectable level has a value of 2 and saturation is at a value of 12, then 2 defi nes what will be represented as black in my image and 12 as white. Between black (2) and white (12) I must divide all of the various shades of gray in my sample into only 9 levels (values 3 to 11). T us subtle diff er- ences in shading will not be captured and stored. On the other hand, if the lowest value is 2 and the highest value is 200, I can capture and store much more subtle changes in light intensity. T erefore, the greater the dynamic range, the better the camera is at capturing small shading diff erences within a specimen. T e dynamic range is the full well capacity of the sensor divided by the read noise. T e dynamic range is oſt en reported as a ratio. A designation of 3,000:1 would indicate a range from 1(black) to 3,000 (white). T e dynamic range can also be reported as how many digital bits of information can be discriminated. A 12-bit detector will have a range of 4096:1 (2 12 = 4096). T e dynamic range will be diff erent at diff erent frame capture rates, so you should compare camera systems at the frame rate you need in your experiments. Dynamic range is another area where CCD systems traditionally exceled, but sCMOS cameras now usually have much better dynamic range. Read noise . Read noise is the inherent electronic noise in the system. T e noise level of the camera is important in determining the dynamic range and also in determining how effi cient the camera will be in low-light situations. Read noise goes up as frame rates increase. T us, although sCMOS cameras are capable of frame rates as high as 100 frames per second, these rates are only useful if you have a very strong signal. For very low-light situations, a modifi cation of the CCD approach called electron-multiplying CCD (EM-CCD) is available. In EM-CCDs the electrons from each photodetector pass through a multi-stage gain register. In each stage of the gain register, multiple electron impact ionization events increase the number of electrons so that the number of output electrons is dramati- cally increased. EM-CCD cameras are thus very sensitive in low-light situations. However, the photodiode size is generally larger than those of traditional cCCDs. Dark noise . Dark noise is that generated by temperature excitation of electrons within the photodetector. Cooled CCDs reduce the dark noise by reducing the temperature. Cooled CCDs exhibit much lower dark noise than equivalent-sized sCMOS sensors. In fact, for most situations the dark noise of cCCD is negligible; not so with sCMOS. However, sCMOS manufacturers in the last few years have greatly reduced the dark noise on their chips, and we may soon see a time when sCMOS dark noise is a negligible contributor to overall readout. Discussion . Microscope camera manufacturers are happy to provide you with a long list of specifi cations for their systems, especially in those areas where their specifi cations exceed their closest competition. However, I have found that the few parameters described above are suffi cient to narrow down the potential list of cameras that would be useful for a specifi c purpose. T en, despite what the specifi cations sheet might indicate, I always compare systems on my microscopes with my specimens to make sure that the camera system is appropriate for my needs. T e practical test is necessary because there is


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Figure 4 : Representative response curve for a CCD photosensor. The sensor has reasonable quantum effi ciency across the full visible spectrum. However, the quantum effi ciency is not equivalent for all wavelengths; the peak effi ciency of 73% is achieved at a wavelength of 560 nm.


chip-to-chip variability and also because each manufacturer includes hardware and soſt ware in their camera that alters the initial photon signal collected by the photodiode. T is signal processing is an integral part of the camera, but that means two cameras from diff erent manufacturers with the same CCD or CMOS chip may have diff erent performance.


Conclusion


Modern CCD and CMOS detectors are the sensors of choice for high-resolution microscope camera systems. Each has its unique features, and deciding which approach best meets your research needs can be daunting. In my experience, the parameters that are the most helpful for deciding if a particular camera system will meet your needs are: pixel size, frame rate, quantum effi ciency, spectral response, dynamic range, and noise. In general, sCMOS systems provide faster acquisition speed and enhanced dynamic range, while cCCD systems off er better low-light sensitivity and more uniformity across the image fi eld for cameras with the same pixel size. EM-CCD can provide even better low-light sensitivity and improved dynamic range compared to traditional cCCD systems but usually have larger pixel size. Once you have sorted out the camera specifi - cations to fi nd potential cameras useful for your applications, always test the cameras under your conditions to make sure the camera is appropriate to your needs.


References [1] J Requejo-Isidro , J Chem Biol 6 ( 3 ) (2013 ) 97 – 120 . [2] A Small , Nature Methods 9 ( 2012 ) 655 – 56 . [3] P Magnan , Nuclear Instruments and Methods in Physics Res A 504 ( 2003 ) 199 – 212 .


[4] T Chen et al. , in Sensors and camera systems for scientifi c, industrial and digital photography applications. Proceedings of SPIE, eds. M Blouke et al., Society of Photographic Instrumentation Engineers, Bellingham, WA, 2000, 451–59.


[5] W Jerome . in Basic Confocal Microscopy, Chapter 7 eds R Price, W Jerome, Springer, New York , (2011 ) 133 – 57 .


www.microscopy-today.com • 2017 September


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