Charles R. Keese

An agglutination assay using emulsified oils

Dispersed fluorocarbon oil droplets were used as the antigen-carrier particle in an immunological agglutination assay. The extent of agglutination was found to vary with the percentage of antigen to that of inert protein used in the solution to coat and stabilize the oil droplets. Maximum sensitivity was observed when 6.25% of the protein adsorbed on the oil droplets was specific antigen. At this percentage, antibodies were detected at concentrations as low as 0.10 micrograms/ml. Results were obtained by visual examination of the reacted samples as well as with quantitative computer analysis using the Zeiss IBAS image analysis system.

Electric Cell-Substrate Impedance Sensing Concept to Commercialization

A personal account of the invention, growth and commercialization of Electric Cell-substrate Impedance Sensing (ECIS) by the inventors of the technology. From the first experiments at the General Electric Research and Development Center in the early 1980s to the outgrowth of applications and finally the incorporation of Applied BioPhysics, Inc., the chapter provides an historical and often amusing account of the evolution of ECIS.

Regiospecific Hydroxylation of Biphenyl and Analogs by Aspergillus parasiticus

The regiospecific conversion of biphenyl to 4,4’-dihydroxybiphenyl (biphenol) by various fungi has been studied for several years. These selective enzymatic hydroxylation syntheses of biphenols are of interest as model xenobiotics for the mammalian metabolism of aromatics (Smith and Rosazza 1974) and as screening systems for carcinogens utilizing 4,4′-dihydroxybiphenyl as the probe (McPherson et al. 1976). Our interest in these systems concerns the production of 4,4’-dihydroxybiphenyl as an intermediate in the plastics and dye industries (Schwartz 1979; Fewson 1981; Cain 1980). For example, the biphenol diol is a monomer in Union Carbide’s polymer Radel™ and GE’s engineering polymer Ultem II™. The organism Aspergillus parasiticus catalyzes the specific conversion of biphenyl to 4,4’-dihydroxybiphenyl in a one-step process.

Attachment and Spreading of Mammalian Cells in Vitro

When mammalian cells are seeded out in tissue culture they will drift down to the surface and firmly attach. The cells attach to a preadsorbed protein layers and not to the naked surface. Normally the protein layer will be a mixture of protein obtained from serum added to the tissue culture medium, but it is also possible to preadsorb pure protein layers onto the surface. This talk will discuss the many factors that affect the interaction of cells with surfaces including the time course of attachment and spreading and the forces that bind the cells to the protein layers.

Enzymes at Oil-Water Interfaces and in Organic Solvents

Enzymes act as remarkably efficient catalysts in biological systems. Some characteristics of enzymes such as their chiral specificity for some substrates make them attractive catalysts for chemical synthesis. Many desirable substrates, however, have poor solubility in the aqueous systems normally employed with most enzymatic reactions. The use of isolated enzymes in organic solvent systems can overcome this problem and offer new possibilities for enzyme use in chemical synthesis. In addition to making substrates with low aqueous solubility available to the enzyme, organic systems can enable enzymatic catalysis at low water concentrations (<0.05%) where new reactions are possible.

Measurement of antigen concentrations with a one-step inhibition assay

A device is described which is used to measure the concentration of antigen in solution. A wedge-shaped gap containing the antigen to be detected is bounded on one side by a reservoir containing specific antiserum and on the opposite side by a detection slide which has been previously coated with a layer of adsorbed antigen. As the antibodies diffuse through the gap, they complex with the soluble antigen in the wedge until these become saturated leaving additional antibody free to complex with antigen on the detection slide. By allowing diffusion to continue for an appropriate time, it is possible to relate the distance along the gap where this has occurred to the concentration of antigen within the gap. Using human IgG as a model antigen, we have been able to measure concentrations down to approximately 10 micrograms/ml with this test.

A Variation of the Radioimmune Assay

A novel method of radioimmune assay is described. It departs from conventional assays in that the radioiodine is reacted with protein following, rather than before, the antigen-antibody reaction. Using a BSA/anti-BSA system, the assay can detect antigen concentrations down to approximately 100 ng/ml.