Luminescent and colorimetric indicators are undoubtedly the most important ingredients of sensor materials. Although they can be used on their own for analyte quantification, for example in microscopic imaging, in this form they represent molecular probes but not sensors. In order to enable reliable quantification of the analyte concentration in various media preparation of a sensor material is necessary. Apart from the indicator such material almost always includes a polymeric matrix (or less commonly an inorganic matrix such as a sol-gel) which acts as solvent and a support for the indicator, but also as a permeation-selective barrier which minimizes the potential interferences. For instance, oxygen indicators are commonly embedded into hydrophobic matrices allowing for free diffusion of oxygen but preventing penetration of polar species (ions, proteins etc.) which can act as luminescence quenchers or alter the environment of the indicator and thus its photophysical properties. Although design of a sensor material might appear trivial from the first glance, it is often far more challenging in reality. First, one should ensure compatibility of the indicator and the matrix, since if poor it would result in dye aggregation and therefore deterioration of the sensing properties. Second, the leaching and migration of the indicator should be prevented. Although non-covalent entrapment in a suitable matrix is often sufficient, only covalent immobilization of the dye can completely eliminate these undesirable effects. It should be kept in mind that migration of the indicator (e.g. into a support) can be significantly accelerated at elevated temperatures (e.g. during autoclavation) whereas leaching may become critical in presence of additional species such as proteins. Third, in order to enable reliable sensing, the materials often contain additional components. These can include: (i) transparent support (typically glass or polyester); (ii) scattering particles (most often titanium dioxide) for enhancement of the sensor signal; (iii) optical isolation layer containing carbon black to minimize inference of the ambient light on the sensor but also the effect of the excitation light on biological systems (particularly for investigation of photosynthesis). Moreover, the sensor materials based on fluorescent indicators commonly contain materials for referencing of fluorescence intensity in ratiometric 2-wavelenght mode or via dual lifetime referencing technique. Once the optimal composition is found, the sensing material can be applied in a variety of formats to create optical sensors (Fig. 1). Most common formats include planar optodes for 2D-imaging applications, fiber optic sensors and microsensors for point measurements, planar sensor spots and nanosensors. Nanoparticle-based sensors have to possess dispensability in aqueous media, low toxicity and cell penetrating properties (if intracellular sensing is desired). Due to the unique property of light to transport multiple information simultaneously, multi-parameter sensors were designed to enable sensing and imaging of several analytes simultaneously with a single sensor. Finally, optical sensing materials can combine sensing functionality with another property, such as e.g. suitability to manipulation with a magnet.
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