PNA openers as a tool for direct quantification of specific targets in duplex DNA.

We report a new approach for target quantification directly within DNA duplex. Our assay is based on the formation of a new biomolecular structure, the PD-loop. The approach takes advantage of a selective hybridization of a probe to double-stranded DNA (dsDNA), which is locally opened by a pair of bis-PNA oligomers. To optimize the technique, several experimental formats are tested with the use of PNA and oligonucleotide probes. The highest sensitivity is achieved when the hybridized probe is extended and multiply labeled with 125I-dCTP by DNA polymerase via strand displacement in the presence of single-strand binding (SSB) protein. In this case, the PNA-assisted probe hybridization combined with the method of multiphoton detection (MPD) allows to monitor sub-attomolar amounts of the HIV-1 target on the background of unrelated DNA at sub-nCi level of radioactivity. The developed robust methodology is highly discriminative to single mutations, thus being of practical use for DNA analysis.

The use of CT in monitoring cryosurgery.

One of the main problems encountered in cryosurgery is that of controlling the amount of tissue that is irreversibly destroyed by cooling. In vitro-experiments in which the homogeneous cooling of various substances, as well as "ice-ball" propagation within different tissues of animals via a cryotip were evaluated. In vivo experiments, on an anaesthetized pig, indicate CT to be a useful continuous-monitoring technique with a high spatial resolution. Using CT simultaneous localization of pathological tissue and measurement of temperature distribution is possible. The density changes induced by cooling were visualized on CT and the corresponding temperature changes were measured using thermocouples. The greatest decrease in density was observed as a change of phase took place. The speed of ice-ball propagation as well as the diameter of the ice ball generated varied considerably depending on the type of tissue. Construction of cryoprobes of a very small diameter could facilitate the percutaneous treatment of lesions within the body.

Contrast in computerized transverse axial tomography of brain.

A study of contrast in photon transmission tomography of brain is presented. Quantitative evaluation of linear attenuation coefficients due to photo-electric, Compton and pair creation effects suggests that the use of photons of a few MeV considerably improves the contrast between white and grey brain matter. The patient dose at different photon energies is calculated.