What’s new in microscopy illumination?

Microscopy illumination systems can often be overlooked when configuring a microscope setup – whether that is for widefield fluorescence or transmitted light applications. However, the type and amount of light hitting the sample plane will make or break an experiment. While microscopy illumination systems have evolved and highly controllable, 8-channel illumination systems are now available, to reach the full potential of this technology it is crucial to optimise not only the illumination but other components such as optical filters. Read on to find out more about the latest developments in microscopy illumination, and which components to consider for bright, high-contrast images and quality data.

CoolLED and microscopy illumination

CoolLED designs and manufactures cutting edge LED microscopy Illumination Systems for researchers and clinicians using the latest LED technology.

Since our team of four introduced the first commercially available LED illumination system for fluorescence microscopy in 2006, we have led the way in transforming microscopy illumination for fluorescence and transmitted applications. Now we are a fast-growing company in Hampshire, UK, with a vast product range and technical expertise spanning optical engineering and the life sciences.

We also understand that something out of the ordinary is sometimes required, and manufacturers wishing to gain a competitive advantage can now access our world-renowned LED technology in customised configurations at www.OEMIllumination.com.

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    Evolution of microscopy illumination


    Before we discuss how to improve microscopy illumination, it is important to first clarify the most popular area of discussion around this topic: intensity and power. As LED microscopy illumination systems have evolved, the topic of optimal microscopy illumination has been surrounded by complex, confusing and sometimes inaccurate measurements and terminology. Even the term ‘intensity’ is inaccurate, and a key issue has so far been an industry-wide lack of a standardised approach to measure illumination. We have set out to change this by introducing a protocol whereby microscopists can quantify the amount of light at the sample plane and therefore directly compare systems to find their ideal solution. We recommend reading the following white paper.

    Evolution of Microscopy Illumination White Paper

    CoolLED’s Microscopy Illumination Product Range

    LED lighting systems are available for fluorescence microscopy and transmitted light applications.
    Take a look at our product range to discover which system is best suited to your requirements.

    Improving microscopy illumination: choosing the right illumination system

    When looking to improve microscopy illumination, the first item to address is the illumination system, and the industry is currently experiencing a transition away from mercury/metal halide towards the LED technology. Since CoolLED introduced the first LED illumination system for fluorescence microscopy in 2006, laboratories have been increasingly adopting this modern light source due to the many benefits:

    • Microscopy Illumination - PerformancePerformance: Irradiance declines over the lifetime of a mercury or metal halide lamp, whereas LEDs are much more stable over time.
    • CostCost: In addition to saving on components, the longer lifetime of LEDs is cheaper in the long-term. Specialist disposal charges due to the high-pressure lamp and mercury content are also avoided.
    • Microscopy Illumination - ControlsControl: LEDs can be easily controlled. They can be switched on and off with precise microsecond timing, which removes the need for a mechanical shutter and improves the temporal resolution of experiments. LED irradiance can also be modulated electronically, removing the need for neutral density filters.
    • ConvenienceConvenience: Lamps frequently need replacing, whereas LEDs are long-lived. The alignment required after frequently replacing lamps is tricky, but LEDs can be factory aligned and are ready to be fitted to the microscope.
    • Safety and the environmentSafety and the environment: Mercury in lamps is hazardous to people and the environment. Lamps are also left on for days during experiments, whereas LEDs can be switched off easily when not in use, which reduces energy consumption. In addition to this, LEDs are more energy efficient and emit much less heat.

    Historically, the irradiance of LEDs was considered low, and although this can still haunt its reputation, the technology has now matured and LEDs are equivalent or brighter in many areas of the spectrum. As the growing efficiency of LEDs leads to irradiance gains, LEDs now typically exhibit above 30% efficiency, whereas traditional lamps are mostly below 10%. As efficiency increases this also reduces heat output, which has an added benefit, since reducing operating temperature is known to increase LED lifetime. Boosting irradiance and extending the lifetime through thermal management is such a significant factor when designing LED Illumination Systems, CoolLED’s work focuses on a range of innovations in this area – for example, reducing LED temperatures through unique heatsink designs which conduct heat away from the LED much more efficiently.

    Microscopy illumination: Choosing the best filters for your application

    Alongside selecting a modern illumination system, microscopy illumination can be increased through optimising other aspects of microscope configuration such as optical filters. Selecting the best optical filters increases irradiance of useful light, since the function of optical filters is to block unwanted light whilst allowing light at favourable wavelengths to pass (i.e. the optimum wavelengths for exciting the selected fluorophore). Filters optimised for a defined set of fluorophores will expose the sample only to the most suitable wavelengths, blocking background noise and leading to brighter and high-contrast images – and better quality data. Historically, filters have been designed for mercury and metal halide lamps and we often come across these being used with new LED microscopy illumination systems, which can compromise image quality, with knock-on effects for data interpretation. Single-band filters are now available that are matched to the spectra of LEDs and popular fluorophores. When used with LED illumination systems, these increase the signal to noise ratio for single-colour imaging and minimise bleed-through to enhance multi-colour imaging. An example of this can be seen below.

    Microscopy Illumination GFP filters

    Try it out!

    If you’re interested in seeing how a modern CoolLED Illumination System can enhance your microscopy illumination, we can provide a free loan unit for testing, with no obligation to purchase. Please contact us at [email protected] for more information.