SIMS microscopy in the biomedical field.

We attempted to indicate the requirements for biomedical applications of SIMS microscopy. Sample preparation methodology should preserve both the structural and the chemical integrity of the tissue. Furthermore, it is often necessary to correlate ionic and light microscope images. This implies a common methodological approach to sample preparation for both microscopes. The use of low or high mass resolution depends on the elements studied and their concentrations. To improve the acquisition and processing of images, digital imaging systems have to be designed and require both ionic and optical image superimposition. However, the images do not accurately reflect element concentration; a relative quantitative approach is possible by measuring secondary ion beam intensity. Using an internal reference element (carbon) and standard curves the results are expressed in micrograms/mg of tissue. Despite their limited lateral resolution (0.5 microns) the actual SIMS microscopes are very suitable for the resolution of biomedical problems posed by action modes and drug localization in human pathology. SIMS microscopy should provide a new tool for metabolic radiotherapy by facilitating dose evaluation. The advent of high lateral resolution SIMS imaging (less than 0.1 microns) should open up new fields in biomedical investigation.     

Adsorption of DNA to mica mediated by divalent counterions: a theoretical and experimental study

The adsorption of DNA molecules onto a flat mica surface is a necessary step to perform atomic force microscopy studies of DNA conformation and observe DNA-protein interactions in physiological environment. However, the phenomenon that pulls DNA molecules onto the surface is still not understood. This is a crucial issue because the DNA/surface interactions could affect the DNA biological functions. In this paper we develop a model that can explain the mechanism of the DNA adsorption onto mica. This model suggests that DNA attraction is due to the sharing of the DNA and mica counterions. The correlations between divalent counterions on both the negatively charged DNA and the mica surface can generate a net attraction force whereas the correlations between monovalent counterions are ineffective in the DNA attraction. DNA binding is then dependent on the fractional surface densities of the divalent and monovalent cations, which can compete for the mica surface and DNA neutralizations. In addition, the attraction can be enhanced when the mica has been pretreated by transition metal cations (Ni(2+), Zn(2+)). Mica pretreatment simultaneously enhances the DNA attraction and reduces the repulsive contribution due to the electrical double-layer force. We also perform end-to-end distance measurement of DNA chains to study the binding strength. The DNA binding strength appears to be constant for a fixed fractional surface density of the divalent cations at low ionic strength (I < 0.1 M) as predicted by the model. However, at higher ionic strength, the binding is weakened by the screening effect of the ions. Then, some equations were derived to describe the binding of a polyelectrolyte onto a charged surface. The electrostatic attraction due to the sharing of counterions is particularly effective if the polyelectrolyte and the surface have nearly the same surface charge density. This characteristic of the attraction force can explain the success of mica for performing single DNA molecule observation by AFM. In addition, we explain how a reversible binding of the DNA molecules can be obtained with a pretreated mica surface.     

UvrD helicase, unlike Rep helicase, dismantles RecA nucleoprotein filaments in Escherichia coli

The roles of UvrD and Rep DNA helicases of Escherichia coli are not yet fully understood. In particular, the reason for rep uvrD double mutant lethality remains obscure. We reported earlier that mutations in recF, recO or recR genes suppress the lethality of uvrD rep, and proposed that an essential activity common to UvrD and Rep is either to participate in the removal of toxic recombination intermediates or to favour the proper progression of replication. Here, we show that UvrD, but not Rep, directly prevents homologous recombination in vivo. In addition to RecFOR, we provide evidence that RecA contributes to toxicity in the rep uvrD mutant. In vitro, UvrD dismantles the RecA nucleoprotein filament, while Rep has only a marginal activity. We conclude that UvrD and Rep do not share a common activity that is essential in vivo: while Rep appears to act at the replication stage, UvrD plays a role of RecA nucleoprotein filament remover. This activity of UvrD is similar to that of the yeast Srs2 helicase.     

Significance of SIMS microscopy for the radioiodine detection in animal and human thyroid tissue.

We defined the SIMS conditions for radioiodine detection in animal and man thyroid follicles, in tissue sections (3 microns) chemically fixed and resin embedded. Two radioisotopes were tested: 125I and 129I, of high (14 mCi 125I micrograms-1) and low specific activity (1.07 10(-6) mCi 129I micrograms-1). In animal study, Wistar rats fed a normal iodine diet (10 micrograms 127I day-1) were injected ip 24 h before sacrifice either with 125I (7 10(-3) micrograms) or with 129I at a dose identical to iodine diet (10 micrograms) or 3 times higher (30 micrograms). No SIMS signal of 125I was obtained in vivo due to its too low concentration, while radioiodine distribution was evidenced with both doses of 129I. Local concentration of previously stored 127I in follicular lumen was not modified, when compared to control (4.14 +/- 0.03 micrograms/mg, m +/- SE), by 125I or 129I at a dose of 10 micrograms, but was nearly doubled with 129I at a dose of 30 micrograms, proof of a pharmacological effect on thyroid iodine regulation. In human study 129I was excluded due to its long half-life (1.6 10(7) years), and 125I was tested only in vitro on two surgical specimens of normal perinodular thyroid tissue maintained in mini-organ culture for 48 h in presence of 100 microCi/ml of 125I. The 125I was detectable, its concentration was 1,000-fold higher than that of 127I (1.5 +/- 0.004 micrograms/mg). For both in vivo and in vitro studies, a positive correlation exists between newly organified radioiodine (125I or 129I) and previously stored iodine (127I).(ABSTRACT TRUNCATED AT 250 WORDS)