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Guidelines for Understanding Magnifi cation in the Modern Digital Microscope Era


James A. DeRose* and Michael Doppler Leica Microsystems , Max Schmidheiny Strasse 201 , Heerbrugg SG , CH-9435 , Switzerland


* James.DeRose@leica-microsystems.com


Abstract: For digital light microscopes, which have image sensors but no eyepieces, the image is displayed directly on an electronic monitor. This development brings a signifi cant change to the usual way magnifi cation of an object viewed via a microscope is determined. Over the years, there have been various standards that defi ne magnifi cation when viewing an image through a microscope’s eyepieces. Only recently have magnifi cation standards been developed that apply to a digital microscope. This article offers guidelines for determining the range of useful magnifi cation values for digital microscopy, particularly when monitors of various sizes are employed.


Keywords: digital light microscopy , magnifi cation range , resolution , megapixel camera , fi eld of view


Introduction


For more than 150 years light microscopy, employing glass lenses to focus light and produce the phenomenon of magnifi cation, has allowed the observation of minuscule entities not visible to the unaided eye. Today there are many types of light microscopes, but here we discuss two of the most common: (1) digital microscopes employing electronic image sensors, but no eyepieces, where the image is displayed on an electronic monitor, and (2) microscopes for direct visual observation which have eyepieces and a digital camera, allowing it to be used in a similar manner to a digital microscope. A feature of interest for any microscope is magnifi cation.


Image magnifi cation is the apparent size of an object at a scale larger (or smaller) than its actual size. Magnifi cation serves a useful purpose only when the resolution of the microscope makes it possible to see more details of an object in the image than when observing the object with the unaided eye. Until recently, magnifi cation has been well defi ned when viewing an image of a sample through the eyepieces of a microscope. For this case, rigorous international standards have been documented [ 1 – 8 ]. Many of these standards also apply to digital microscopy, but strict defi nitions and standards for magnifi cation achieved by a digital microscope, where the image is viewed on an electronic monitor, have been developed only in the last few years [ 9 ]. In research publications and presentations, images must include a calibrated scale bar or a statement of the width of the fi eld of view, typically in nm or µm. However, when viewing images in real-time on monitors of various sizes, indications of scale may not be shown, and the instantaneous magnifi cation of an image may not be known. Moreover, there is typically a lower and higher limit of magnifi cation which defi ne the range of useful magnifi cation . Digital microscopes, as well as microscopes for visual observation equipped with digital cameras, allow the rapid acquisition of high-quality images. T ese instruments are applied in various scientifi c and engineering fi elds. T e goal of this article is to provide digital microscopy users with helpful guidelines so they can determine the range of useful magnifi cation when making observations and measurements. T e specifi c parameter values given here are


20


Of course, the upper limit of visual magnifi cation depends on the maximum resolving power of the microscope system. When the magnifi cation is pushed above the useful range, no additional details about the sample can be seen. T is situation is referred to as empty magnifi cation [ 10 ]. Even within the


for instruments from Leica Microsystems, but the principles described are quite general for all light microscopes employing digital image acquisition and display.


Magnifi cation


What exactly is magnifi cation? Magnifi cation is the ratio of the size of an object in an image produced by an optical system to the actual size of the object itself. T us, lateral magnifi cation, M DIS , can be defi ned as:


Figure 1 : (a) Stereo microscope (M205 C) equipped with a digital camera. The ant sample can be observed via the eyepieces or on an electronic monitor displaying the image from the sensor in the camera. (b) Digital microscope (DMS1000) shown with two different monitor sizes for image display.


doi: 10.1017/S1551929518000688 www.microscopy-today.com • 2018 July


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