The fluorophore Fura-2 has an absorption spectrum that varies markedly depending on the concentration of calcium (Ca2+) that is present near the fluorophore molecule. By measuring the ratio of intensities on digital images captured using two different excitation wavelengths, the variation of calcium concentration as a function of location on the sample can be tracked.

The conventional approach to ratiometric imaging with Fura-2 is based on exchanging two excitation filters in a filter wheel. Now with Semrock’s VersaChrome TBP01-380/16 tunable excitation filter, ratiometric imaging with Fura-2 can be more carefully optimized to your specific experimental conditions. This filter makes it possible to monitor the Fura-2 signal corresponding to virtually any excitation wavelength between 340 and 380 nm – continuous tuning like a monochromator, yet with the high transmission, steep edges, and high out-of-band blocking available only with optical filters. And, wavelength tuning of a VersaChrome filter can be 2 to 3 times faster than exchanging filters with even the highest-speed filter wheel.

The added flexibility of a tunable excitation source can even enable new types of experiments. For example, the same excitation filter used for high-contrast ratiometric imaging can be tuned to excite Fura-2 at its isosbestic point, thus enabling monitoring of the Fura-2 signal independent of calcium concentration.