This page contains a Flash digital edition of a book.
Microscope Illumination: LEDs are the Future


James Beacher CoolLED, CIL House, Charlton Road, Andover, SP10 3JL, UK jim.beacher@coolled.com


Introduction Light microscopes in laboratories and hospitals are used for


examining many different types of samples—from industrial research to life-science research and clinical screening. Tese procedures use conventional bright-field, differential phase contrast (DIC), and fluorescence microscopy among other techniques. In all cases, the light source on the microscope has a crucial influence on the quality of images viewed and the conclusions reached.


Microscope Illumination For low-magnification macroscopic illumination, a single


point-light source is oſten used. Because it is not in the optical path, the illumination is at an oblique angle. Tis oſten may be adequate, but shadowing can be a problem for applications where even illumination is required. In order to overcome shadowing and poor homogeneity, it is common to use multiple light sources or some form of ring-light positioned around the objective lens to generate an even illumination. Recently, fiber optics have been used for low-magnification illumination; light is delivered to the microscope via a fiber bundle from a remote lamp. In high-magnification light microscopy applications, the optimum method of sample illumination is by delivering the light through the optical path of the microscope itself. In this case, there are many additional parameters to consider, as described below. Historically, illumination for samples in light microscopy


has been provided by a range of conventional incandescent or discharge lamps [1]. Tey can be classified as: halogen, tungsten- halogen, xenon, mercury (also known as an “HBO” or a “UV Burner”), and metal-halide. In recent years, light-emitting diode (LED) illumination products have become available that offer many exciting benefits. In this article, the different types of illumination available are reviewed. Te benefits and disadvantages of each type will be identified and compared with LED-based products for performance, convenience, safety, environmental effects, and operating costs.


Incandescent and Discharge Lamps If you are using a microscope in your lab, it is likely that


it will have illumination fitted using one of the conventional lamps. Te most common type of microscope illuminator is the tungsten incandescent lamp. Tese lamps are relatively inefficient and can exhibit a shiſt in color temperature with time [1]. Most lamps supplied today are of the more advanced tungsten-halogen or quartz-halogen type. Tese generate light across the visible spectrum, but much of the energy is dissipated as heat in the infrared (IR) region, and there is little illumination below 400 nm. Tey are rarely used for fluorescence microscopy, which needs higher intensities. Xenon lamps exhibit a flatter intensity across the spectrum than halogen, which makes them more suitable for quantitative analysis.


18 For high-magnification and fluorescence microscopy,


the high-pressure mercury vapor arc-discharge lamp is most common. Tese are significantly more powerful than other forms of lamp, but their intensity varies across the spectrum. Tey are hampered by poor spatial homogeneity because of their complex construction and alignment requirements. Replacement bulbs can be difficult to align, leading to uneven illumination over the microscope’s field of view. Bulb lifetime is a few hundred hours. Te metal halide lamp is an enhanced version of the


high-pressure mercury lamp. Tese more expensive bulbs provide better illumination stability and have a longer bulb lifetime. However, they still suffer from degradation in performance during the bulb lifetime (around 2,000 hours). Because they produce significant amounts of UV light, a liquid light-guide used to deliver light from the illumination unit to the microscope ages and needs to be replaced regularly [2]. Benefits. All these conventional lamps have the benefit


that they produce a broad spectrum of white light that can be used for many applications. In fluorescence microscopy, the user simply changes the microscope filter cube to match the fluorophore being used; common fluorophores used are DAPI, GFP, FITC, etc. [3]. Disadvantages. Conventional lamps have a number of


significant drawbacks. Tey generate unwanted heat, are inefficient, and require the regular replacement of bulbs. Mercury-based lamps can be dangerous for the user because they generate considerable UV light, which can damage the eye. Conventional lamps produce UV light, which is typically not required for illuminating the sample. In fact, this UV light can bleach samples, killing live tissue cells and reducing the amount of time that a sample can be examined. Although it is rare, these bulbs can explode and release toxic mercury vapor into the laboratory air. A period of warm-up and cool-down is required before


and aſter use, which is inconvenient. As a result, these lamps are oſten kept switched on all day in order to be available when required. Because they are inefficient, energy is wasted and unwanted heat is generated. Te bulbs have a limited operating life (typically measured


in hundreds of hours) and degrade in performance over that period [1]. Intensity from a conventional lamp decreases through its life (Figure 1). Most lamps are quoted with a lifetime to 50% of original intensity. Tis means that the illumination of a sample varies dramatically through time. Any quantitative or comparative measurements cannot be done reliably unless a new bulb is used on every occasion.


LED Illumination With LED-based microscopy illumination, almost all of


the disadvantages of conventional incandescent and discharge lamps can be overcome. LEDs are solid-state semiconductor


doi:10.1017/S1551929511000411 www.microscopy-today.com • 2011 July


Page 1  |  Page 2  |  Page 3  |  Page 4  |  Page 5  |  Page 6  |  Page 7  |  Page 8  |  Page 9  |  Page 10  |  Page 11  |  Page 12  |  Page 13  |  Page 14  |  Page 15  |  Page 16  |  Page 17  |  Page 18  |  Page 19  |  Page 20  |  Page 21  |  Page 22  |  Page 23  |  Page 24  |  Page 25  |  Page 26  |  Page 27  |  Page 28  |  Page 29  |  Page 30  |  Page 31  |  Page 32  |  Page 33  |  Page 34  |  Page 35  |  Page 36  |  Page 37  |  Page 38  |  Page 39  |  Page 40  |  Page 41  |  Page 42  |  Page 43  |  Page 44  |  Page 45  |  Page 46  |  Page 47  |  Page 48  |  Page 49  |  Page 50  |  Page 51  |  Page 52  |  Page 53  |  Page 54  |  Page 55  |  Page 56  |  Page 57  |  Page 58  |  Page 59  |  Page 60  |  Page 61  |  Page 62  |  Page 63  |  Page 64  |  Page 65  |  Page 66  |  Page 67  |  Page 68  |  Page 69  |  Page 70  |  Page 71  |  Page 72  |  Page 73  |  Page 74  |  Page 75  |  Page 76  |  Page 77  |  Page 78  |  Page 79  |  Page 80  |  Page 81  |  Page 82  |  Page 83  |  Page 84