Microscopy101
An Image is Everything: A Tutorial on Choosing and Using the Components of a Dynamic Data Capture System
Duncan Stacey* and Robert Gurney Linkam Scientific Instruments Ltd *
duncanstacey@linkam.co.uk
Abstract: Dynamic imaging—acquiring images and simultaneously recording relevant sensor parameters—is a powerful way to extend the information and insight available over and above optical micros- copy. It improves understanding and correlates how properties of materials alter with, for example, changing temperature, pressure, humidity, tensile forces, shear stress, or aging. The basics of an imag- ing setup are the microscope, a relevant stage, a camera of appro- priate performance, and suitable software to acquire, synchronize, and analyze the images and data collected. However, many factors need to be considered and optimized to ensure reliable experiments. These include basic features of the microscope—objective resolution and magnification, for example, as well as the accuracy and repro- ducibility of the stage, the camera format, dynamic range, and reso- lution. In this tutorial, we discuss the components of a dynamic data and image capture system and present examples where researchers have used this approach to better understand their materials.
Keywords: Dynamic imaging, image capture, thermal microscopy, sCMOS cameras, resolution
Introduction Tey say “a picture is worth a thousand words.” For
materials changing under a range of environmental condi- tions, this certainly holds true. Dynamic imaging—acquiring images and simultaneously recording relevant sensor param- eters—is a powerful way to understand and correlate how the properties of materials alter with, for example, changing temperature, pressure, humidity, tensile forces, shear stress, or aging. Tis oſten requires an integrated system capable of recording and imaging dynamic processes that occur during an experiment. Many materials change color, shape, and size with chang- ing conditions. For example,
ferroelectric materials realign
(Figure 1), and thermotropic liquid crystals undergo phase transitions and color changes, and self-assemble (Figure 2) as temperature changes; polymeric films can tear when under ten- sile strain (Figure 3); and many materials change color as they oxidize at high temperatures. Of course, optical microscopes visualize these changes but, in more and more applications, microscopists now need to capture images with a specialized camera in order to correlate them with stage sensor parameters to either quantify a transformation or gain additional infor- mation about visual differences. Events such as a color shiſt or structural change can be analyzed, the speed of formation and extent of a tear can be measured, and the size and shape of particles can be evaluated.
44 doi:10.1017/S1551929520001558
Assembling a Dynamic Data and Image Capture System Recent advances in technology have led to the avail-
ability of many new products that are ideal for microscopy. With a wide range of options, it is important to choose the microscope system, including the stage and image capture configuration, best suited for dynamic research applications and sample characteristics. Te basics of an imaging setup are straightforward. System components are comprised of the microscope, relevant
stage (to accurately and repro-
ducibly control temperature, environmental conditions, or tensile/shear forces), CCD or CMOS camera of appropriate performance, and suitable soſtware to acquire, synchronize, and analyze the images and data collected. Some of the latest advances, such as scientific CMOS (sCMOS) cameras, have focused on offering high sensitivity and speed, which is oſten ideal for live cell fluorescence imaging. However, they are not necessarily the best option for many material science appli- cations where dynamic samples evolve on the microscale, especially with changing environmental conditions such as temperature and humidity. However, like many laboratory setups, the devil is in the details. Questions to consider include:
• Is my microscope optimized—objective resolution and magnification, numerical aperture?
• How accurate and reproducible is my stage? • What camera format is best matched to my microscope? • What camera resolution is needed for the application? • How do chip size, dynamic range, and pixel size and spac- ing affect results?
• Which magnification should be used to get the best image? Moreover, if video images are to be captured, parameters
such as frames per second and time-lapse ability must also be considered and optimized.
Selection of an Appropriate Camera Selecting a camera with the appropriate performance
for microscopy can suffer from the same biases seen in the consumer camera world. More is oſten simply considered better—particularly when it comes to sensor size and pixel count—without due regard for the need to, for example, bal- ance the improved spatial resolution of smaller pixels with
www.microscopy-today.com • 2020 November
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