Resonance energy transfer (RET) in hybrid polymer/inorganic/organic layers in saponite films, prepared using the ‘layer-by-layer’ (LBL) method, was studied. This method allows the fabrication of a multicomposite molecular assembly based on electrostatic and/or other intermolecular interactions between the interacting species. The LBL films were designed by sequential deposition including positively charged polycations ( poly(diallyldimethyl ammonium), PDDA) and negatively charged saponite dispersions with donor molecules (rhodamine 6G) and saponite dispersions with acceptor molecules (rhodamine 3B). In this sequence, the layer of saponite donor molecules was placed on an adjacent layer of saponite acceptor molecules, while the layers were separated by inserting a PDDA polycationic layer. The adsorption process was studied using absorption spectroscopy. The linear deposition regime was completed when ten layers had been deposited. The process of REF was studied by fluorescence spectroscopy. In this case the donor molecules – rhodamine 6G – play the role of lightharvesting antennae and transfer the energy to the next layer covered by energy acceptor, i.e. rhodamine 3B molecules. The insertion of polycation layers is rationalized by the adhesion of negatively charged underlying layers and the stabilization of multicomponent systems. The energy transfer efficiency was affected heavily by changing the distances between the adjacent layers.

The ability to observe the behavior of living cells and tissues provides unparalleled access to information regarding the organization and dynamics of complex cellular structures. While great strides have been made over the past 30 to 40 years in the design and application of a variety of novel optical microscopic techniques, until recently, it has not been possible to image biological phenomena that occur over very short time periods (nanosecond to millisecond) or over short distances (10 to 1000 Å). However, the recent combination of (1) very rapidly gated and sensitive image intensifiers and (2) the ability to deliver fluorescence excitation energy to intact living biological specimens in a pulsed or sinusoidally modulated fashion has allowed such measurements to become a reality through the imaging of the lifetimes of fluorescent molecules. This capability has resulted in the ability to observe the dynamic organization and interaction of cellular components on a spatial and temporal scale previously not possible using other microscopic techniques. This paper discusses the implementation of a fluorescence lifetime imaging microscope (FLIM) and provides a review of some of the applications of such an instrument. These include measurements of receptor topography and subunit interactions using fluorescence resonance energy transfer (FRET), fluorescence anisotropy of phospholipids in cell membranes, cytosolic free calcium (Ca2+)i and the detection of human papillomavirus (HPV) infection in clinical cervicovaginal smears.