Invasion Assay techniques

In summary, chemiluminescence is a sensitive, non-invasive technique that can measure reactive oxidant production by small numbers of neutrophils indeed, neutrophil-derived chemiluminescence can be detected in as little as 5 fA of unfractionated human blood. The assay is suitable for automation using either multichannel luminometers or luminescence microtitre plate readers. Many researchers, however, have questioned the usefulness of this technique because of the uncertainty of the nature of the oxidant(s) that are detected. Nevertheless, in view of the recent developments made towards the identification of the oxidants measured and the assay s ability to detect intracellular oxidant production, it is has an important place in the phagocyte research laboratory. 

Disulfoton induced the liver MFO system in animals (Stevens et al. 1973). In the same study, exposure to disulfoton orally for 3 days also increased ethylmorphine N-demethylase and NADPH oxidase activities, but had no effect on NADPH cytochrome c reductase. Thus, the induction of the MFO system required repeated dosing with relatively high doses. Furthermore, these changes are not specific for disulfoton exposure, and these subtle liver effects require invasive techniques in humans to obtain liver tissue for performance of these enzyme assays. 

The measurement of compounds in bioprocesses, including fermentations, using conventional laboratory techniques such as HPLC, TLC or calorimetric assays is often tedious, invasive, requires sample handling and difficult to do in real time. For a bioprocess where it is important to gain information about the reactor status for feedback control, methods enabling rapid and reliable measurement of components are desirable. 

It is important to note that the fluorescence and light scattering assays are non-invasive techniques and thus can be coupled to other techniques such as biochemical fractionation or electron microscopy. The sample size and concentration of reagents are in the same range as those used in other techniques, thus facilitating comparison. 

From Equation (17) an analysis can be made if the quantity of ML can be determined as a function of ligand added at each aliquot. However considerable errors may arise as this quantity must be assayed, after each aliquot of ligand is added. However non-invasive techniques such as isothermal microcalorimetry calorimetry can be used to directly determine the quantity of ML without disturbing the system. For a calorimetric analysis an estimation of the enthalpy change for the macromolecule-ligand interaction must be made. An assumption can be made that at the start of the study, the initial aliquot of ligand added to the system, if sufficiently small, will completely bind to the macromolecule (as initially the macromolecule will be in excess). The enthalpy change associated with this interaction can then be used to calculate the A// for the interaction (Equation (3)). 

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