the liquid the optical field does not end abruptly at the reflective interface but its intensity falls off exponentially with distance this surface electromagnetic field called the u201c evanescent wave u201d can selectively excite fluorescent molecules in the liquid near the interface the thin evanescent optical field at the interface provides low background and facilitates the detection of single molecules with high signal to noise ratio at visible wavelengths the evanescent field also can image fluorescently labeled nucleotides upon their incorporation into the attached template primer complex in the presence of a polymerase total internal reflectance fluorescence microscopy is then used to visualize the attached template primer duplex and or the incorporated nucleotides with single molecule resolution nucleic acid template molecules include deoxyribonucleic acid dna and or ribonucleic acid rna nucleic acid template molecules can be isolated from a biological sample containing a variety of other components such as proteins lipids and non template nucleic acids nucleic acid template molecules can be obtained from any cellular material obtained from an animal plant bacterium fungus or any other cellular organism biological samples of the present invention include viral particles or preparations nucleic acid template molecules may be obtained directly from an organism or from a biological sample obtained from an organism e g from blood urine cerebrospinal fluid.