Calcium imaging Light Source optimised for Fura-2: CoolLED pE-340fura
The CoolLED pE-340fura is a bespoke LED Illuminator for Fura-2 ratiometric calcium imaging. The Light Source allows high-precision, stable, high-throughput imaging with video-rate time resolution. Resulting from our collaboration with the University of Strathclyde, the pE-340fura utilises the successful pE-300 Series platform, and also supports everyday fluorescence microscopy in a compact and affordable package.1
The CoolLED pE-340fura includes 340 nm and 380 nm LEDs for ratiometric calcium imaging with Fura-2. Adding flexibility to this calcium imaging Light Source, this also includes white light for everyday fluorescence microscopy.
Ultimate control of your calcium imaging light source
Control is where LEDs come into their own as a calcium imaging light source. Precise temporal control is crucial, as slower recordings reduce experimental accuracy. The solid-state nature of LED calcium imaging light sources means LEDs have fast and precise on/off switching. As with any peripheral device, latencies can be introduced by USB serial communications and computer operating system overheads. This can reduce switching speeds by as much as 100 ms and may also impact system synchronisation, e.g. between the camera and calcium imaging light source.
For users wishing to capitalise on the maximum speed of LEDs, electronic control via BNC TTL offers switching speeds of around 20 μs. The trade-off here is cost, and TTL control often requires specialist computer cards and components which can be expensive.
To overcome this and make TTL a cost-effective means of achieving high-speed imaging, the CoolLED pE-340fura LED Illumination System includes the Sequence Runner program. Sequence Runner uses the TTL output signal available on most scientific cameras and cycles though LEDs in a user-selected sequence for each TTL signal. This is a simple approach and provides high-speed microsecond switching speeds that are precisely synchronised to the camera exposure.
Improved signal-to-noise
Work by Sandrine Prost et al., from the University of Edinburgh, has shown that with independent wavelength controllable LED sources, signal-to-noise is dramatically improved over bulb systems and even over some available white broad-spectrum LED sources.
High levels of autofluorescence and fast photobleaching of specific fluorescence when illuminating Qdots with Metal Halide.2
Preventing phototoxicity and photobleaching with your calcium imaging Light Source
When combining the narrow spectral spread of each wavelength with TTL control, this limits unnecessary sample irradiance, minimising phototoxicity and photobleaching. Helping to reduce these effects, it is also possible to modulate the brightness of the LED calcium imaging Light Source to optimally excite the Fura-2 dye. The irradiance of LED Illumination Systems has been increasing year-on-year, and modern systems feature simple and precise irradiance modulation in 1% steps (0-100%), which can be controlled from software, analogue connection or the control pod.
With the resulting improvement in cell viability, this achieves not only insightful but accurate data. It also reduces costs since less Fura-2 is required per experiment.
