Standardization of the chromium-51 release, cell-mediated cytotoxicity assay: cryopreservation of mouse effector and target cells

Utilizing controlled cryopreservation techniques, we were able to standardize the 51Cr release cytotoxicity assay and thereby ensured reliable comparisons between results obtained on different days. Optimal conditions for freezing of both effector and target cells were quite similar. Dimethyl sulfoxide (DMSO) at a concentration of 7.5-10.0% was employed as the cryoprotective agent and cells were frozen at the rate of -1 degrees C/minute. The handling procedures for the cells before and after freezing were important. Factors affecting recovery of functional reactivity were related to toxicity of DMSO for the cells, the osmotic stress placed upon the cells as the DMSO was being removed after thawing, the handling temperature of the freshly thawed cells, and the susceptibility of cells to mechanical damage immediately after thawing. The recovery of lymphocytes after freezing was about 70%; the recovery of cytotoxicity was around 85%. Syngeneic cytotoxic reactivity induced by inoculation with the Moloney strain of murine sarcoma virus was cryopreserved, as were allogeneic cytotoxicity and natural cytotoxic reactivity. Multiple tests employing effector cells from the same frozen pool gave reproducible results; the standard error of the mean percent cytotoxicity was less than 1.5%. Cryopreserved target cells gave decreased day-to-day variability in susceptibility to lysis, since the same population of cells could be employed in each assay. These results demonstrated conclusively that we can now have a constant source of effector cells and target cells, which can be used from assay to assay as an internal standard.     

Complementary energy methods in the computation of electrostatic fields

It is shown how complementary energy methods can be applied to problems involving the calculation of electrostatic fields. The results obtained using such methods provide error bounded solutions to the system energy, and may be compared with those found through application of the conventional finite element method. To illustrate the technique the classical problem of a cylindrical tank, half filled with a charged liquid, is investigated. Comparisons are made on the basis of computer C.P.U. time, and core storage requirements, for a given degree of accuracy in total system energy. The question of local error in primary field quantities is also examined.     

Triple-deck solutions for subsonic flow past hymps,steps, concave or convex corners and wedged trailing edges

The application of two distinct numerical approaches to the titled problems is discussed. One is the recently developed fast scheme due to Veldman; the other involves techniques to speed up the earlier schemes by faster but accurate boundary layer routines and mixed updating. Both are second order accurate and give virtually identical results. Simple transformations are used to cover completely the infinite range of streamwise intergration, and reversed flow is treated iteratively. Separated flow solutions are presented and discussed.     

Arrhythmias during halothane anaesthesia II: the influence of atropine

The effect of i.v. atropine premedication on cardiac rhythm was studied in healthy adult patients during thiopental-N2O/O2-halothane anesthesia without intubation. A higher incidnece of arrhythmias was seen in younger patients in close relation to administration of atropine, but the overall incidence during anesthesia was identical in atropine groups and the control groups. The most common arrhythmias were supraventricular ectopies. None of the ECG irregularities led to serious arrhythmias. No consistent changes in blood pressure were observed as the result of arrhythmias or changes in heart rate. It is concluded that atropine should be reserved for situations where severe bradycardia and hypotension occur, or can be expected to occur, and not given automatically, since cardioacceleration which is inherent in its action may be injurious to patients with limited cardiac reserve.     

The preparation and polymerization of ethylene-d4

A direct method for preparing deuterated ethylene by reducing acetylene-d₂ with chromous ions in a dimethylformamide/deuterium oxide mixture is described. This method gives a good yield of precursor gas of relatively high purity. The gas was polymerized with a Ziegler-type catalyst to produce deuterated polyethylene with sufficient purity and melt flow properties for use in laser fusion studies. A reaction variables study showed that polymerization of deuterated ethylene yields polymers with higher molecular weights than those obtained using normal ethylene. The study also indicated that the deuterium content of the polymer decreases with increasing reaction temperature, thus dictating that the process operates near room temperature. Several mechanical and chemical properties of the deuterated polymer were determined and are reported.     

Barley grain as carbon substrate for fungal protein production in a farm process

A simple, non-aseptic, submerged aerobic fermentation was sought in which microbial protein could be synthesised from non-protein nitrogen, at the expense of barley carbohydrate, to supplement the protein content of barley grain for non-ruminant feeding. Screening of 23 isolates of fungi in shake flask cultures showed that microbial protein could be successfully produced by a number of different species in a medium containing barley as a source of carbohydrate. Protein synthesis was markedly affected by the addition of different sources of non-protein N to the fermentation; and (NH₄)₂SO₄ or urea proved to be good N sources in this respect. Aspergillus oryzae was selected as the most suitable species for protein synthesis, based on its performance when supplemented with the various N sources. The growth rate of A. oryzae in stirred culture was such that high yields of mycelial protein could be achieved within 24 h from a culture inoculated with a spore suspension. Under the conditions of growth used in these experiments there appeared to be little advantage in adding more than 2% barley to the medium. Higher yields of protein could be achieved at barley concentrations in excess of 2%, but only if the incubation period was extended. The tolerance of A. oryzae to low pH values was exploited to enable fungal cultures to be grown under non-aseptic conditions. Good yields of protein were obtained in 40 litre cultures grown at pH 3.5 in a modified domestic washing machine. Although protein yields were poor, non-aseptic growth was also sucesssful at pH 3.5 in 1000 litre cultures grown in an agricultural feed-mixing vessel.