Search

Three types of methods for bichromatic paper disk spectrophotometry are described and illustrated by the detailed presentation of procedures for the dosage of hemoglobin and protein (with some applications in virology), as well as for glucose determination. The general algorithms that would make possible the automation of these methods by means of programmable systems are presented.

A linear approximated model is presented that takes into account a local anisotropic distribution of scattered light within a turbid medium. Light intensity depends upon the distance from the boundary surface of the scattering centers, so that the probability of photon-chromophore interaction is a function of the distance. For the sake of simplicity it is assumed that there are only two main zones, corresponding to two probabilities of photon-chromophore interaction. In this form, the Kubelka-Munk theory fits the experimental data supplied by bichromatic photometry of nucleic acids on paper disks.

The bichromatic paper disk spectrophotometry ultramicromethod developed in the "Stefan S. Nicolau" Institute of Virology can be successfully applied to the study of viral and cellular nucleic acids. The main applications - succinctly presented in the review - refer to : the identification and spectral characterization of nucleic acids, their quantitative determination, the study of the helix - statistical coil transition, the evaluation of the degree of polymerization of single stranded polynucleotides, the study of the kinetics of chemical and enzymatic hydrolysis and the determination of RNase activity, e.g. in human serum, the demonstration of the interaction between nucleic acids and other molecules of biological interest (biopolymers, poly- and oligocations, etc.), the rapid characterization of the ion exchange capacity of chromatography papers and of a mobile phase of the water - alcohol - electrolyte type.

Bichromatic absorbance measurements on RNA-impregnated paper disks may be applied to the ultramicroanalytic evaluation of the ion exchange capacity (IEC) of different chromatography papers. A newly defined absorbance parameter and the curves of its variation in terms of IEC are used to this purpose.

Bichromatic spectrophotometry determinations were performed on Whatman chromatographic paper disks impregnated with rRNA solutions of different concentrations, at ionic strengths ranging from nearly 0 to 1.4 M. The disks consisted of cellulose, either native or modified (by grafting of carboxymethyl, phosphate, Ecteola or DEAE groups, or by loading with silica gel). The results demonstrate that: 1) there is a positive correlation between the absorbance enhancement by multiple light scattering and the nearness of the nucleic acid molecules in relation to the surface of cellulose fibrils; b) the action on absorbance of ionic strength and of other physical factors differs in terms of the electric charges and of the exposure of other groups that may interact with nucleic acids through hydrogen bonds or Van der Waals forces.

A bichromatic spectrophotometry study in the near UV spectral range was performed on paper disks impregnated with DNA and RNA in NaCl solutions of different concentrations. Absorbance curves plotted versus a NaCl molarity log-scale degree of polymerization and the solubility of the polynucleotide. The -a/b value, determined by a previously reported equation, appeared to be constant for a given nucleic acid, irrespective of the wavelength in the spectral range 205-280 nm. This phenomenon cannot be attributed to a heterogeneous macroscopic distribution of the nucleic acid throughout the disk.

The results of over 200 scientific papers in the field of in vivo and in vitro virus--tumor relationships, worked out in the "Stefan S. Nicolau" Institute of Virology during a period of over 25 years, are very briefly reviewed. Stress is laid on the original theoretical and practical contributions of the institute to the knowledge of viral carcinogenesis in animals and humans. Some personal viewpoints in the interpretation of results are presented and new research prospects in this field--which represents a traditional concern of the institute--are lutlined.

RNA absorbances Ad were determined by bichromatic spectrophotometry at 240, 260 and 280 nm, versus 350 nm values, on paper disks containing different 2-propanol concentrations and NaCl amounts. No significant differences in terms of the NaCl content could be observed at alcohol concentrations higher than 70%; the maximum Ad value was found to correspond to about 50% 2-propanol. The possible phenomena implied in the absorbance changes are discussed taking into account the peculiar action of 2-propanol on the water structure and content in the bulk phase within the paper capillaries and inside the macromolecular and aggregation structures of RNA and cellulose.

Studies in human embryo fibroblasts infected with measles or herpes simplex virus showed a reduction in virus yield when cultures were pretreated with 1-10 mM lithium chloride doses. Maximum effect was obtained by a 1 h treatment with 10 mM lithium chloride, preceding viral infection by 19-24 hours. A specific antiviral effect against measles virus was manifest immediately after culture pretreatment. Intermittent treatment with 10 mM lithium chloride of cultures persistently infected with measles or herpes virus obtained from human myeloid K-562 cell line shows a reduction in the extracellular virus yield. In the K-562/herpes virus system, the culture treatment with lithium chloride and acyclovir (10 microM) has an additive inhibitory effect on virus production. The paper is focused on the mechanism of lithium chloride antiviral action and the expediency of lithium therapy in SSPE (subacute sclerosing panencephalitis).

A bichromatic ultramicromethod on paper disks was used to determine the age-dependence of capillary blood RNase activity in healthy infants and young children.

The paper reports on serological results obtained by HAI tests against several alphaviruses - eastern equine encephalitis (EEE), western equine encephalitis (WEE), Chikungunya, Midlebourg, Semliki Forest, Sindbis, and Ilheus viruses - carried out in humans and some domestic animals in a region with an intensive circulation of migratory birds. It was possible to detect in human antibodies to EEE, WEE, and Sindbis viruses, but in very low proportions (1.8%, 0.4%, and 0.6%, respectively). Serological results obtained in animals were practically negative: antibodies to EEE virus were found only in one out of 93 horses investigated.

Bichromatic paper disk spectrophotometry allowed the demonstration of an analogy between the increase in the solubility of highly polymerized RNA complexed with streptomycin cations (which accounts for the preservation in vivo of the infectivity of a viral RNA--streptomycin complex) and the effect of a cellulose/solution interface on the distribution of the highly polymerized RNA.

The paper is a review of researches done at the "Stefan S. Nicolau" Institute of Virology, in Bucharest, the last five years, in the antiviral chemotherapy field, as well as of the chemotherapeutical implications of studies conducted about other aspects of virology.

The report is a review of the research and results of the "Stefan S. Nicolau" Institute of Virology scientists in the field of herpes viruses, as they were materialized in the great number of published or communicated scientific papers these last ten years.