Pilot-scale verification of a computer-based simulation for fractional protein precipitation

The development and experimental verification at pilot scale of a suite of models for the batch precipitation by two-cut ammonium sulphate salting-out of total protein and alcohol dehydrogenase from yeast homogenate is presented. The model consists of two elements: protein and enzyme solubility prediction and precipitate phase particle property prediction. An isotherm-based approach has been used successfully to describe solubility behaviour for a range of operating conditions typical of those obtained at a process scale. Estimation of the precipitate phase particle size distributions has been achieved through a discretized population balance approach using simplified terms to account for particle breakage and aggregation. The developed model accounts for the effects of average shear rate and residence time in the precipitation vessel across a two orders of magnitude range of scale. The size-density relationship for the precipitate phase has been defined. Results of simulations are compared with pilot scale verification data to confirm the validity of the models developed.     

Exercise-induced muscle damage, repair, and adaptation in humans

This study examined exercise-induced muscle damage, repair, and rapid adaptation. Eight college-age women performed three eccentric exercises of the forearm flexors. One arm performed 70 maximal contractions (70-MAX condition), and the other arm performed 24 maximal contractions (24-MAX) followed 2 wk later by 70 maximal contractions (70-MAX2). Criterion measures of serum creatine kinase, muscle soreness and pain, isometric strength, and muscle shortening were assessed before, immediately after, and for 5 days after each exercise. Significant changes in all criterion measures were found after the 70-MAX exercise with a slow recovery that was not complete by day 5 after exercise. The 24-MAX condition showed only small changes in the criterion measures. Changes in the criterion measures after the 70-MAX2 exercise were significantly smaller than those after the 70-MAX exercise. Results from this study, with regard to the ability of the muscle to adapt to exercise-induced damage, suggest that an adaptation takes place such that the muscle is more resistant to damage and any damage that does occur is repaired at a faster rate. It is also clear that a relatively small insult will produce this adaptation.     

INACCURACY OF ESTIMATIONS OF S PHASE FRACTION BY REDUCTION IN CLONING EFFICIENCY WITH HYDROXYUREA OR TRITIATED THYMIDINE

The current hypothesis, that the fractional reduction of cloning efficiency in semi-solid culture systems induced by pretreatment of the cells with hydroxyurea (HU) or [³H]TdR equals the fraction of cells initially in S phase, is tested. A lymphoblastoid cell line, SK-L7, with known cell cycle kinetics was exposed to cytotoxic concentrations of HU or suicidal doses of [³H]TdR and then initiated in semi-solid and liquid culture. Although approximately 0.6 of the initial population was in S, 1-hr exposures of HU at concentrations of up to 10^-2 M failed to reduce subsequent cloning efficiency. the 1-hr exposure to HU did not reduce either the immediate cell number or the gross population doubling rate over 24 hr. A 24-hr exposure to 10^-3 M HU reduced the cloning efficiency by approximately 98%, confirming the drug's cytotoxic capability. [³H]TdR at doses of 100 μCi/ml for 20–40 min reduced the cloning efficiency by approximately 60 and 70%, respectively. Although no cytotoxicity immediately after exposure was observed in either case, gross population doubling rate in liquid culture was reduced. While HU failed to reduce subsequent cloning efficiency, [³H]TdR reduced cloning efficiency by approximately the fraction of initial cells in S. the above hypothesis, therefore, cannot be applied naïvely as a technique for quantitating the fraction of a clonogenic cell population in S phase.     

A comparative investigation of soluble protein polymorphism in Robinia pseudoacacia roots by polyacrylamide gel electrophoresis

Investigations of soluble proteins by polyacrylamide gel electrophoresis of root extracts of black locust (Robinia pseudoacacia L.) were carried out with 41 trees from diverse habitats representing dominant-stem forms (R. p. var. rectissima Raber) and typical forms (R. pseudoacacia L.). Soluble protein patterns of dominantstem forms and typical trees did not show differences attributable to tree form. Heritability estimates (broad sense) were determined as 9·19% within location and 7·.5% among populations. A variance components model was constructed which showed the interaction between parental trees and location to be most significant in determining variation. Location variance was second in importance, with parental variance and experimental error of less significance. The data were analyzed by the moment of inertia. It is indicated that, based on protein similarity, the dominant-stem form is an ecological variant and should not be given varietal status.     

Discrepancy between flow cytometric analyses of CALLA using two monoclonal antibodies

Both the J5 and BA-3 monoclonal antibodies are considered to be specific for epitopes on the common acute lymphoblastic leukemia antigen (CALLA). Flow-cytometric analyses of three cell lines and one normal bone marrow sample using these antibodies as CALLA markers demonstrated that J5-labeled cells were always brighter than those labeled with BA-3, and that the ratio of their fluorescence intensities varied widely in the different systems. Furthermore, one of the lines, RPMI 8226, while positive for J5, appeared to be negative when labeled with BA-3, except for a slight displacement of the fluorescence distribution relative to the control. A possible explanation for the observed results is that the BA-3 binding epitope or epitopes on CALLA may vary in their number and/or accessibility to the antibody. These observations suggest that the use of a single monoclonal antibody to detect a cell surface antigen may be misleading, particularly when a negative result is obtained.