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Biomedical subjects

D B Roodyn

Publications and source records attributed to D B Roodyn.

9 recordsLinked to original sources
PubMed1970

A generalized system for automation of enzyme assays.

A flow system was developed, using a Technicon AutoAnalyzer, that is readily adaptable to a range of enzyme assays. The system includes lines for pumping substrate, cofactor, buffer and enzyme and for generating linear gradients. By using a variable-speed proportioning pump the incubation time may be continuously varied, and the system also allows for continuous variation in the pH, substrate or cofactor concentration, incubation temperature and enzyme concentration. A FORTRAN V program was written that uses instrument calibrations to calculate the flow rates in the individual lines, the incubation time and the characteristics of the gradient used. The computer then prints out instructions for preparation of reagents to give a required reaction mixture, weighing sheets for stock solutions and the results of the assay in international units in suitable tables and graphs. The flow system and computer program are designed to facilitate the automation of manual assays. A detailed example is given of the use of the system [the assay of three dehydrogenases in yeast: l(+)-lactate dehydrogenase, d(-)-lactate dehydrogenase and succinate dehydrogenase], and the general applications of the method are discussed. The program has been deposited as Supplementary Publication no. SUP 50002 at the National Library for Science and Technology, Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies may be obtained on the terms indicated in Biochem. J. (1970), 116, 7.

Autoanalysis↗
PubMed1965

Further study of factors affecting amino acid incorporation into protein by isolated mitochondria.

1. The incorporation of amino acids into protein in isolated mitochondria has now been studied in more detail, and with mitochondria from a range of tissues. 2. Liver mitochondria from newborn rats are twice as active as those from adult rats. 3. The mitochondria are inactivated by excessive homogenization and repeated freezing and thawing. 4. The incorporation is sensitive to conditions of incubation and in particular to the rate of oxygenation, shape of vessel and depth of fluid. Best results are obtained by incubation in flat-bottomed vessels containing suspensions with less than 3mm. depth of fluid. 5. The requirement for oxidizable substrates has been examined with a range of substances, and most of the common energy-yielding metabolites of the mitochondrion are effective. Their activity is greatly influenced by concentration, and some, but not all, of the substrates show optimum concentrations for incorporation with decreased activity at higher concentrations. 6. Some amino acids can act as energy sources for the incorporation. 7. The effect of increasing the concentration of labelled amino acid is different for different amino acids, and complex effects occur on the addition of amino acid mixtures. 8. It is concluded that the amino acid incorporation into protein finally obtained is the result of many interacting factors related to the structural and metabolic state of the mitochondrion.

Amino Acids↗