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The work presented in this thesis was initiated in order to develop a non-invasive real-time gas phase analytical technique, based on selected ion flow tube mass spectrometry (SIFT-MS), for monitoring the progression of human cell cultures and the detection of microbial contamination in such cultures by monitoring and quantifying the emitted volatile compounds.\ud \ud Fundamental SIFT experiments were performed to characterise the reactions of the SIFT-MS precursor ions (H3O+, NO+ and O2 +●) with several volatile compounds of potential value to biological research; a necessity for their quantification. The work has resulted in new methods for the quantification acetaldehyde and CO2 in gaseous samples.\ud \ud The compounds present in the headspace of sealed cultures of six human cell types were analysed by SIFT-MS, the key finding being their consumption of the toxic volatile compound acetaldehyde from the media. Further experiments involved the addition of the enzyme aldehyde dehydrogenase inhibitors diethylaminobenzaldehyde and disulfiram to cultures of hepG2 (hepatocellular carcinoma) cells, when it was observed that consumption of acetaldehyde from the cultures/headspace was reduced, and in some cases, acetaldehyde was even produced due to the actions of the cellular alcohol dehydrogenase enzyme. Furthermore, the solvent dimethylsulphoxide was reduced to imethylsulphide by the cells, which is known to occur via the enzyme methionine sulphoxide reductase. This process was retarded by the ALDH inhibitors.\ud \ud The use of SIFT-MS for the detection of microbial infection in mammalian cell cultures was also explored. The volatile compounds emitted by E. coli (strain JM109), into the gas phase above two different culture media, were analysed using SIFT-MS. Further, the progression of a culture of this bacterium was monitored continuously over a 4-hour period. The findings of this research were then applied to the study of human cell cultures intentionally infected by E. coli bacterium, including cultures contained in a 1L bioreactor.

Lymphocyte survival changes observed at 1, 2 and 3 days as responses to 3 doses of ionising radiation in vitro (40, 100, and 500 cGy) are analysed by computer according to a simple (single cell population) mathematical model. Intrinsic radiosensitivity, the susceptibility to lethal injury, which is expressed as the D 37 value (the radiation dose permitting 37% survival), is estimated separately from the kinetics of subsequent death of lethally-irradiated cells (expressed as their half-life, or t 1 2 value). Among the 35 patients with B-cell CLL studied (15 were never treated), both parameters varied widely and independently of one another, t 1 2 ranged from 9–200 h and above, D 37 from 14–500 cGy or above. Twenty-three patients were deemed ‘radiosensitive’ ( D 37 below 110 cGy). D 37 level did not correlate with treatment status, mode of treatment, clinical staging (Rai) or lymphocyte count. With some exceptions, D 37 remained relatively constant for individual patients with increasing duration of disease or alterations in treatment status. The assay method may prove useful as an aid in predicting response to low-dose splenic irradiation (SI) in CLL.

On exposure to the phorbol ester 12-O-tetradecanoyl-13-acetate (TPA) the pathological (non-dividing) lymphocytes of B-cell chronic lymphocytic leukaemia (CLL) lose their characteristic ultrasensitivity to the cytocidal action of colchicine in vitro. They are no longer killed in 1 day by the drug at 10−6 M-concentration. The effect was the same whether the cells were incubated in the continuous presence of TPA, or subjected instead to pulse-treatment with it (for as little as 5 min.). Colchicine at one thousand times greater concentration was now needed to kill the cells. CLL lymphocytes already primed to undergo interphase death by pretreatment with colchicine could be prevented from doing so by early addition of TPA. A marked proportion of those CLL lymphocytes destined to undergo early spontaneous death in vitro in the absence of colchicine could be prevented from doing so by TPA. The loss of colchicine ultrasensitivity applied to cells which had not yet undergone TPA-induced morphological transformation to blast-like cells or differentiation to cells containing abundant cytoplasmic immunoglobulins (Clg). These transformed cells materialised in greatest incidence (70–80%) after 3 days of culture, an observation in agreement with others workers.