AbstractRecent advances in thin film optical coating technology significantly improve the filters available for fluorescence spectroscopy. Bandpass and long‐ and shortpass filters with very sharply defined edges can provide from 10−5to 10−6blocking within 10–15 nm of the transmission region and are ideal for use as excitation and emission filters. A variety of nonpolarizing dichroic beamsplitters for use in epi‐illumination configurations or in multiple emission configurations provides optimum longpass, shortpass, band reflection, or bandpass spectral control. These dichroics, used with high‐performance bandpass, longpass, or shortpass filters, form matched sets that optimize the signal‐to‐noise ratio and system efficiency for fluorescence spectroscopic systems in single or multiple dye applications. Specially designed dichroic beamsplitters are used to reduce excitation filter overheating. Other dichroic beamsplitters efficiently separate two planes of polarization in a narrow wavelength band. Rejection band filters can be used to measure the fluorescent dye Indo 1 with very low e