One of the most crucial components of your microscope is the light source. Correct illumination allows for optimal sample observation, picture capture, and data analysis. There are various approaches to improve your system depending on the type of fluorescence excitation light source (mercury arc lamp, mercury metal-halide lamp, or LED). This post will provide you with some pointers on how to improve your fluorescence light source.
Traditional mercury arc lamps are normally housed in a lamp house attached directly to the microscope, though their output may be transmitted via a light guide on systems like confocal microscopes. Follow these four best practices while following the manufacturer's recommendations to extend the life of these light sources:
Reduce the number of times you turn the lamp on and off:
Do you need to leave the room for a few moments? Keep the light turned on! A lamp's life is shortened by frequent on/off switching. Although certain mercury lamp power units contain a cooldown waiting mechanism to prevent repeated switching, being aware of this feature might help you increase the life of the bulb.
Check to see if your bulb is still in good condition:
After only 70 hours, the intensity of mercury bulbs begins to fade. They are substantially weaker after 200 hours. Make sure you replace your bulb according to the manufacturer's recommended lifespan (200 or 300 hours). Always follow your company's hazardous waste policy when disposing of these bulbs.
Align the bulb as follows:
Homogeneous field illumination is the outcome of proper alignment. Review your user handbook to make sure you're doing things the right way with your lamp house. Placing a white paper card on the microscope stage is an easy way to see if your bulb is misaligned. Then, using blue excitation, examine it at low magnification. Because cellulose fluoresces spontaneously, uneven illumination will be visible.
Keep a backup on hand:
In the event of a bulb failure, always have a spare on hand. It's not uncommon for old mercury bulbs to quit working, and you don't want to be caught without one when you're ready to image your samples. Remember to replace the bulb according to the manufacturer's instructions (usually after 200 or 300 hours, depending on the bulb) and dispose of the old one as hazardous waste containing mercury.
A pre-centered long-life metal-halide bulb doped with mercury is used in mercury and metal-halide lamp systems. These bulbs are pre-centered in the main power unit and typically last 2,000 hours. Although there are a few direct-mounted units on the market, illumination is normally given to the microscope via a liquid light guide (LLG).
For the first 1,000 hours, such light sources produce a pretty steady output, after which the intensity begins to diminish. For spectrum output over time and replacement procedures, consult the manufacturer's technical information. As with mercury bulbs, you should dispose of the old ones in accordance with your company's hazardous waste policy.
Many people are unaware that the LLG deteriorates with time and becomes much less effective after 4,000 to 6,000 hours of use. Use the built-in shutter on the main power unit instead of the one on the microscope to block excitation light. This will aid in the LLG's efficiency being maintained for a longer period.
Nonetheless, following the second or third bulb replacement, you should plan to replace the liquid light guide. Liquid light guides do not need to be centered, but they must be fully installed in the power unit socket as well as the LLG adapter on the microscope. When changing the LLG, make sure to follow the manufacturer's recommendations.
Many advantages exist for upgrading from a mercury-based fluorescence light source to a light-emitting diode (LED) unit in terms of microscope performance and simplicity of maintenance, including:
Mercury has more and brighter spectral peaks.Without the need for a warm-up period, the switch can be turned on and off instantly. Exceptionally long life span (20,000 hours or more) There's no need to buy or align bulbs (yay!). Because of the lower energy use and the absence of mercury, it is environmentally friendly. At important wavelengths like 488, 568, and 647, several LED sources deliver peak spectral output brighter than mercury. Models with three to 16 distinct, rapid-switching excitation wavelengths are offered.
LED sources have quick on/off timings and most have varying intensities, some of which are controlled by wavelength. They can be triggered through USB or a transistor-transistor logic (TTL) signal from your PC/software to shutter wavelength. Individually triggered wavelengths are ideal for converting an older manual microscope to an automated multichannel fluorescence microscope (using a multichannel filter set). Convert your old manual inverted microscope into a live-cell time-lapse imaging system by adding a Z-focus device, an environmental chamber, and necessary software.
Because LED systems don't require any alignment and don't have any bulbs to replace, they're a lot easier to manage. They endure tens of thousands of hours, emit no mercury, and use a fraction of the energy. As a result, they can broaden the range of applications that your older microscope can handle while lowering the overall cost of ownership.
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