Characterization of glyoxalase I purified from pig erythrocytes by affinity chromatography

The linear order of nine fragments generated by the action of endonuclease AvaI on the DNA of bacteriophage lambda was determined from the altered fragmentation patterns of bacteriophages containing known deletions and of hybrids of bacteriophages lambda and phi80. Digestion of 5'-terminally 32P-labelled bacteriophage-lambda DNA was used to identify the terminal fragments. Measurement of relative fragment lengths permitted rough mapping of the endonuclease-AvaI cleavage sites relative to the ends of the bacteriophage-lambda chromosome. The fragment order was confirmed and the map refined by analysis of the fragmentation of derivative phages containing single cleavage sites for endonuclease EcoRI.     

Calcium phosphate transfection generates mammalian recombinant cell lines with higher specific productivity than polyfection

Transfection with polyethylenimine (PEI) was evaluated as a method for the generation of recombinant Chinese hamster ovary (CHO DG44) cell lines by direct comparison with calcium phosphate-DNA coprecipitation (CaPO4) using both green fluorescent protein (GFP) and a monoclonal antibody as reporter proteins. Following transfection with a GFP expression vector, the proportion of GFP-positive cells as determined by flow cytometry was fourfold higher for the PEI transfection as compared to the CaPO4 transfection. However, the mean level of transient GFP expression for the cells with the highest level of fluorescence was twofold greater for the CaPO4 transfection. Fluorescence in situ hybridization on metaphase chromosomes from pools of cells grown under selective pressure demonstrated that plasmid integration always occurred at a single site regardless of the transfection method. Importantly, the copy number of integrated plasmids was measurably higher in cells transfected with CaPO4. The efficiency of recombinant cell line recovery under selective pressure was fivefold higher following PEI transfection, but the average specific productivity of a recombinant antibody was about twofold higher for the CaPO4-derived cell lines. Nevertheless, no difference between the two transfection methods was observed in terms of the stability of protein production. These results demonstrated the feasibility of generating recombinant CHO-derived cell lines by PEI transfection. However, this method appeared inferior to CaPO4 transfection with regard to the specific productivity of the recovered cell lines.     

Percutaneous nephrostomy--a retrospective study focused on palliative indications

We present an overview of the fast development of less invasive techniques in intrarenal surgery all based on percutaneous nephrostomy. Life-long urinary diversion with nephrostomy is often necessary in patients with malignant disease and such patients have more postnephrostomy complications than patients with kidney stones and their survival is short. In a follow-up of 246 patients with 275 nephrostomies performed consecutively over two years, mean survival after urinary diversion was only 7.9 months in 38 prostate cancer patients with ureteral obstruction and only 5.3 months in 20 patients with advanced bladder cancer. We emphasize the necessity of informing the patient and his/her family of the expected outcome of the procedure and the importance of using carefully chosen and realistic indications.     

Affinity of putrescine,spermidine and spermine for pigmented tissues

Determinations of the amounts of putrescine, spermidine and spermine in tissues from mice and cows indicated that the eye melanocytes are very rich in these substances. The concentration of these di-and polyamines was also found to be much higher in pigmented than in albino hair of mice. The melanin polymer has the character of a polyanion — explaining the affinity of these cations for pigmented tissues. Further experiments indicated that these substances to a considerable extent may reach the melanin-containing tissues via the circulation.     

Glyoxalase I, a zinc metalloenzyme of mammals and yeast.

Glyoxalase I from human and porcine erythrocytes, rat liver, and yeast was shown to contain zinc in a stoichiometry of about 1 mole per mole of enzyme subunit. Removal of zinc by means of a chelator caused elimination of the catalytic activity. The mammalian apoenzymes could be partially reactivated by addition of Zn²⁺.