It’s fair to say that when it comes to light sources for fluorescence microscopy, the amount of light reaching the sample plane is a huge concern when choosing, comparing or optimising systems. After all, fluorescence demands a lot of light to excite the fluorophore and still make sure there’s enough emitted energy left for the sensor to provide bright, beautiful images – and quality data.
For all its importance, the industry has not made up its mind on how to measure or even discuss this topic with consistency. Some discuss power, others discuss intensity. We think it’s time to set the record straight, so let’s start with what these terms really mean:
- Power: Measured in W, this is largely meaningless without considering how effectively power is focused onto the sample plane. You should also question where in the system the measurements have been taken. If anywhere other than the sample plane (i.e. LED chip or end of a liquid light guide), remember that light is lost as it travels through to the sample and these numbers will be inaccurate.
- Intensity: Power per unit area. It can refer to any kind of power (electrical, heat, light etc.) and is measured in W/m2. This is easy to confuse with radiant intensity which measures power per unit steradian (i.e. power per unit cone angle of emission) in W/sr.
There is a term which refers to the light power at a surface per unit area, measured in mW/mm2. This is irradiance and we use it to describe the amount of light at the sample plane, which is the most meaningful place to measure light.
Now we have settled on appropriate terminology and the most accurate place to measure irradiance, we have also put forward a protocol for standardised measurement, which can be found in our white paper.
The success of fluorescence microscopy hinges on the light source, and we have set out to increase the confidence of microscopists when discussing, comparing or working to improve irradiance.