The aim of this study is to investigate whether aberrant peripheral blood lymphocytes can be corrected by the addition of a highly diluted quinone to the culture medium. The oxygen uptake of both normal and aberrant lymphocytes is examined before and after the quinone is added. The results will indicate whether the quinone is able to increase the oxygen uptake rate of aberrant lymphocytes, thereby returning them to normal metabolism.

 It is hypothesized that the addition of this highly diluted quinone to a solution of aberrant lymphocytes will have a beneficial effect on the blood cells by boosting their immune response and returning them to normal metabolism. This reaction would result in an increase in the level of oxygen uptake within the solution which can be monitored polarigraphically. Materials and Methods The main apparatus in this experiment is the Digital Model 10 Oxygen Electrode (Rank Brothers Ltd, Cambridge, UK) used to measure the amount of dissolved O2 in a solution. The temperature in the chamber of the oxygen electrode is controlled by a DC10-P5 water bath (ThermoHaake, UK) and kept at a constant 37˚C throughout the experiment. The data from the electrode is logged using a Delta Logger (Delta-T Devices, Cambridge, UK) which has also been programmed to take continuous measurements of room temperature, water bath temperature, and the temperature at the base of the water chamber on the electrode where the stirring mechanism is located, using separate SKTS 200/U temperature probes (Skye Instruments Ltd, Powys, UK). Humidity and room temperature are logged using a SKH 2011 probe (Skye Instruments Ltd, Powys, UK). The level of light is recorded using an LX-01 light meter (Lutron). The Delta Logger is programmed and controlled by a PC using Ls2Win 1.0 software, collecting readings from each of these probes and devices every second, then calculates and records the mean average over each 5 sec. period. Peripheral blood lymphocytes are isolated in a nutrient medium (RPMI 1640) and standardized. 

Initially readings are taken for about an hour, to allow any temperature effects to settle down and to ascertain the rate of O2 uptake by the cells. A 0.5ml solution containing 10-6 diluted parabenzoquinone is then added to the test chamber and readings are monitored for further hour to examine the effect of the quinone on the rate of O2 uptake in the solution. The electrode uses a magnetic stirrer in the chamber to circulate the solution, so it is possible that the magnetic field from this may have an adverse effect on the experiment. To mitigate this, a thin layer of polythene is inserted into the chamber to separate the stirrer from the reaction. Initial tests to validate the equipment and procedures involved have been carried out using E. coli bacteria cultivated in RPMI-1640 solution. The bacteria have during the three month pretest period digested all but a trace of the glucose in the solution, which makes them ideal to use as a model. O2 uptake from the bacteria can be measured before and after provision of additional glucose. Results The work is ongoing at present; however a sample of the results collected is given below. Figure 1 shows how the voltage collected from the oxygen electrode, which is proportional to the partial pressure of oxygen in the medium under test varies over time. Used in Fig.1 is 2ml of an RPMI-1640 solution containing E. coli bacteria, the bacteria had consumed all but a trace amount of glucose in the original container.

 Half way through the 2 hrs approx. experiment, at 11:27am 0.5ml of a 0.05% glucose solution was added to feed the bacteria. The subsequent initial increase is due to the dissolved oxygen in the glucose solution. The voltage then drops steadily due to the bacteria feeding on the glucose, since their increased metabolism leads to a fall in the amount of oxygen present in the test sample. Figure 2 shows the results from a similar experiment performed as a control. The initial test solution is 2ml of distilled water. As before, half way through at 1:43pm, 0.5ml of a 0.05% glucose solution is added. It is clear that both before and after the glucose was added the level of dissolved O2 remains the same. As mentioned above, the apparatus was designed to allow continuous measurement of not only the level of dissolved oxygen, but also room temperature, humidity, the light level, water bath temperature, and the temperature at the base of the water chamber on the electrode. Examples of this data can be seen in the figures below.  Half way through the 2 hrs approx. experiment, at 11:27am 0.5ml of a 0.05% glucose solution was added to feed the bacteria. The subsequent initial increase is due to the dissolved oxygen in the glucose solution. The voltage then drops steadily due to the bacteria feeding on the glucose, since their increased metabolism leads to a fall in the amount of oxygen present in the test sample. Figure 2 shows the results from a similar experiment performed as a control. The initial test solution is 2ml of distilled water. As before, half way through at 1:43pm, 0.5ml of a 0.05% glucose solution is added. It is clear that both before and after the glucose was added the level of dissolved O2 remains the same. 

As mentioned above, the apparatus was designed to allow continuous measurement of not only the level of dissolved oxygen, but also room temperature, humidity, the light level, water bath temperature, and the temperature at the base of the water chamber on the electrode. Examples of this data can be seen in the figures below. cells however is that they do not so readily uptake dissolved O2. The rate at which the dissolved O2 is used up can be examined using the procedure described above, to evaluate the normality of the cells. The aim of this study is to identify if highly diluted parabenzoquinone can correct the metabolic function of aberrant lymphocytes. The introduction of parabenzoquinone, a high redox potential quinone into separate culture mediums of normal and aberrant lymphocytes will allow us to examine if the addition of the quinone affects the rate of oxygen uptake in the samples. An increase in oxygen consumption may indicate that the cancer cells are returning to normal and re-establishing their original metabolic pathway. The initial results from experiments using E. coli confirm the sensitivity of the system; since there is a clear distinction between test samples with and without bacteria when adding glucose. Acknowledgement The financial assistance of Children with Leukaemia and the additional helpful suggestions of Dr Bob Brown of Glamorgan University’s Applied Sciences Dept is gratefully acknowledged for this work.