APUD cell development in grafts from chick or quail presumptive enteric endoderm combined with a fragment of notochord : E.L.M.J. Wiertz-Hoessels and J. Drukker. Department of Anatomy/Embryology, Medical Faculty, University of Limburg, Maastricht, the Netherlands

Abstract

Characterization of the internal environment of a sol–gel matrix is an important area of investigation in optical biosensors. In the present study, different sol–gel compositions were prepared by varying the water (H2O) to tetraethyl-orthosilicate (TEOS) ratio (R) from 1 to 16 and the changes in the internal environment of the sol–gel both in bulk and thin films as a function of aging (storage) were investigated using fluorescence spectroscopy. We focussed on the fluorescence characteristics , viz. emission and excited state lifetime of Hoechst 33258 (H258), a bisbenzimidazole derivative, which was used as fluorescence probe entrapped in the TEOS derived sol–gel bulk and thin films. These sols were prepared at a low pH (∼2.0) and the thin films were coated by dip coating technique at withdrawal speeds of 1 cm/min and 0.1 cm/min. Usually, uniform thin films were obtained at a high speed (1 cm/min) and partially cracked film at a low speed (0.1 cm/min) as observed by fluorescence microscope. These observations did not change during aging. On the contrary, three months long observations on steady-state fluorescence emission measurements on H258 depicted a blue shift from 535 nm to 508 nm at R = 1 in the sol–gel bulk, whereas at higher ratios this was not prominent. At all ratios, dual emission bands were observed in thin films. This may be due to faster sol–gel to xerogel transition during aging depending on the ratio (R). Analysis of the excited state decay profiles of H258 revealed a double exponential fitting having a short (τ1) and a long (τ2) component in both fresh and during aging, in the sol–gel bulk and thin films, indicating heterogeneity in the internal environment. The value of τ1 increased from 0.4 ns to 1.2 ns whereas τ2 attained a value from 3.0 ns to 3.6 ns at R = 1 upon aging in the sol–gel bulk. The corresponding values of τ1 and τ2 in thin films were 0.3 ns and 3.5 ns, respectively. The values of these decay components in thin films did not alter much due to storage, but their relative contributions showed more systematic changes in the thin films. The observed changes could be correlated to rigidification in the bulk depending on the ratio (R). This process was very slow at R ≥ 4. The heterogeneity in the internal environment of bulk and thin films upon aging appeared to be different as revealed from analysis of excited-state lifetime. Thus, the bisbenzimidazole derivative H258 appears to be very useful probe for characterizing the internal environment of both the sol–gel bulk and thin films.