Cairn Research Limited unveils a joint project with University of Oxford, Dept. of Pharmacology, to develop a novel TIRF system

As well as our Max Planck collaboration on the PAM development, we are also running a joint project with Dr. Nigel Emptage at the Department of Pharmacology in Oxford, for the development of a Total Internal Reflection Fluorescence (TIRF) system with some novel features.

Most of the TIRF systems supplied by the microscope manufacturers send the excitation light via the objective, which must therefore have a very high numerical aperture objectives in order to allow the light to exit from it at a sufficiently high angle to achieve TIRF criteria. Such objectives tend to be expensive and difficult to use, with the highest-performance ones requiring special immersion oils and coverslips as well. We have chosen to commercialise the alternative approach of using the upper surface of a prism as the TIRF interface, and viewing the fluorescence from above through whatever objective the user prefers. Another advantage of this approach is that it readily allows the angle of incidence of the excitation light to be varied over a broad range.

This is potentially very useful, as the depth to which the excitation light penetrates from the prism into the sample varies with the angle of incidence. In our implementation, we use a galvanometer drive (similar to the ones used in our Optoscan monochromator) to deflect the excitation light beam over a wide range on a millisecond timescale. This allows sequential images to be taken at different excitation angles, and hence with the TIRF effect penetrating to different depths. By seeing how the relative fluorescence of particular structures varies with angle, we can gain information not only on their relative position, but also on whether and if so how quickly they are moving towards or away from the interface.

This information is particularly useful for studies of exocytosis or endocytosis, as a preliminary report from our collaboration (as shown in our poster presentation displayed at the Physiological Society Meeting in Bristol, July 2005) has already demonstrated. This system is now going into production, so we are in a position to take orders. The cost of a basic system is 10,000 Euros. This includes a USB interface and control software. You will also require a suitable light source, which is usually a laser, but an LED may also be bright enough for some applications (we can advise on request). For an additional 3,000 Euros a second galvo drive and ancillary optical components can be fitted, allowing simultaneous and independent control of excitation at two different wavelengths.