Current Pharmaceutical Biotechnology - Volume 5, Issue 3, 2004

The collection of articles published under the motto “The way down from single genes and proteins to single molecules” is covering the results of groundbreaking developments of the recent years. They include single molecule detection yielding ultimate sensitivity and closely linked to the analysis of molecular noise by correlation spectroscopy. Several important examples from the field of nucleic acid as well as protein ana Read More

The present review gives a short summary on techniques useful for single molecule research, describes experiments on in vitro single molecule detection and reactions of single molecules and finally reports on the behavior of single molecules and single virus particles in living cells. One experiment on single molecule enzyme kinetics of lactate dehydrogenase, an enzyme used in the diagnosis of heart attacks and one ex Read More

Protein conformational dynamics, often associated with static and dynamic inhomogeneities, plays a crucial role in biomolecular functions. It is extremely difficult to characterize such inhomogeneous dynamics in an ensembleaveraged measurement, especially when the protein involves in a multiple-step complex chemical reaction, such as an enzymatic reaction. Single-molecule spectroscopy is a powerful approach to analyz Read More

Recently developed single-molecule spectroscopy (SMS) permits the analysis of fluorescent mixtures one molecule at a time. SMS methods provide the means to make rapid measurements on small, complex samples without the need for separations and target amplification enabling a new class of ultrasensitive nucleic acid assays. Here we give a brief overview of the current state of the art of SMS nucleic acid analysis and discus Read More

We have devised a new technique based on single fluorescent molecule detection for the analysis of specific sequences of DNA. The method consists of synthesizing a fluorescent reporter molecule using a polymerase extension reaction and labeled nucleotides. The fluorescent reporter products are analyzed in a laser-based single-molecule detection system. We have applied this method to the detection of pUC19 and B Read More

This mini-review describes how single-molecule sensitive fluorescence resonance energy transfer (FRET) and photoinduced electron transfer (PET) reactions can be successfully applied to monitor conformational dynamics in biopolymers. Single-pair FRET experiments are ideally suited to study conformational dynamics occurring on the nanometer scale, e.g. during protein folding or unfolding. In contrast, conformational dynami Read More

We present novel technical features and results from a two channel confocal fluorescence lifetime microscope, which allows to efficiently investigate fluorescence dynamics down to the single molecule level. The MicroTime 200 time-resolved fluorescence microscope offers a multicolor excitation where different picosecond diode lasers are used. For imaging and positioning purposes we utilize a compact Piezo scanner w Read More

Recent advances in the development of new microscopical techniques with single-molecule sensitivity have given access to essentially new types of information on biological systems. In this review, basic methodological concepts of ultra-sensitive microscopy are presented and characterized, with focus on their applicability for a bioanalytical instrument. Measurements on artificial lipid bilayers were used to evaluate the feasibili Read More