Glucose uptake assay

Time and concentration dependence of glucose uptake and degradation by Simian virus modified (i.e. adipocyte-like) mice fibroblasts, stimulated by the galactosylpicolinato complex 10b (Figure 5.3).The ordinate indicates the absorption (in arbitrary units) in an MTT assay as a measure of the amount of glucose degraded in the cells after uptake. 

The above assays (glucose uptake and oxidation by isolated rat adipocytes hexose transport by muscle cells and activation of the insulin receptor tyrosine kinase) are among the standard methods for the in vitro screening for potential antidiabetic drugs (498, 499). The previous examples show that interpretation of the results of such tests for metal complexes should commence with verification of the stability of the complex (or speciation of the metal ion) in the reaction medium. 

Phloretin is the aglycon of phlorizin and inhibits the facilitated diffusion of glucose catalyzed by GLUT1 or GLUT4. It has been used to terminate the uptake of glucose in timed assays with isolated membranes or reconstituted transporters. 

Some enzyme-catalysed reactions result in the production or uptake of a gas and some of the earliest assay methods used this phenomenon as a basis for monitoring the reaction. Classically, Krebs, in elucidating the metabolism of glucose, used such methods. They measure either the pressure changes that... 

Figure 10. Production of malondialdehyde, the change in Chlorella viability and uptake of O3 by a suspension of Chlorella cells. A sample from a culture of 38°C grown C. sorokiniana uflr. pacificensis (3 X10 cells/ml autotrophic medium) was treated with 180 ppm ozone. Thiobarbituric acid reactants were assayed by the method of Heath b- Packer (23), viable cells by plating on glucose-supplemented agar medium, and ozone uptake on a Cary spectrophotometer as described in Figures 6-8.

Binding of antibody to T4-E decreases enzyme activity. The unbound T4-enzyme conjugate, on the other hand, is active and catalyzes the oxidation of glucose-6-phosphate with simultaneous reduction of NAD-t to NADH. The rate of increase of absorbance at 340 nm is inversely proportional to the number of unsaturated TBP binding sites. Test results are expressed as percent of T uptake using a calibration curve or a mathematical function. Manual and automated versions of this assay are commercially available. ... 

Morris et al. (495) observed a Cr(III) dependent increase in 2-deoxyglucose uptake by cultured mouse myotubes in the presence of insulin. The effect was caused by the addition of nanomolar concentrations of CrCl3 to the reaction medium [a standard minimal essential cell culture medium, containing amino acids, mineral salts, and glucose, which was purified with Chelex resin to remove trace heavy metal ions before the addition of Cr(III)]. The same Cr free medium was used for cell growth [growing the cells in the presence of a trace amount of Cr(III) reduced their sensitivity in the assays]. Thus, unlike the previous studies (491), an apparent insulin-potentiating effect of CrCls was demonstrated. However, Cr(III) most likely formed complexes with amino acids of the medium (80), which prevented its hydrolysis in a neutral aqueous solution and made it more available to the cells. 

Figure 1. Time course of o-glucose and uptake from the growth medium and increase in dry weight of mycelia of P. charlesii cultures, Penicillium charlesii was cultured in a modified Raulin-Thom medium (24). The contents of the flasks were removed at intervals, filtered and the filtrate was assayed for (O—O) total carbohydrate, (A—A) dry weight, and (A—A) pPGM (23). The initial concentrations...

Figure 4.9. Effects of chloramphenicol on the oxidation of -alkanes and glucose by propionic acid bacteria. Strains P. pentosaceum CCM (A, B) P.coccoides (C). Oxygen uptake was assayed in the presence (a) or absence (b) of chloramphenicol at 100 pg/ml. Substrates glucose (1,2) w-alkanes (3,4). From Vorobjeva et al. (1979a).

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