Possible Hydrogen-Bond interactions of steroid hormones-II. corticosteroids

For each of the polar functions of the twenty known crystalline structures of corticosteroids, we determine the position of the atoms forming an intermolecular hydrogen bond with the oxygen of the function, so that we can find the preferential positions-relative to the steroid-of the atoms that take part in such bonds. That leads us to an estimate of the distance between the atoms capable of fixing the steroid by O₃ and O₂₀ or O₂₁ on the receptor. The determined value is 16.5 Å. We apply the same reasoning to explain the role of the inhibitors.     

A simple spectrophotometric method for measurement of nematode populations.

A convenient automated method for measuring inorganic phosphate based on the malachite green reaction with a phosphomolybdate complex has been developed. Less than 100 pmol of inorganic phosphate can be readily quantitated by the method which utilizes standard AutoAnalyzer equipment. Inorganic phosphate is measured in sample volumes of less than 0.1 ml and without interference by a number of phosphorylated metabolic intermediates or nucleotides. This methodology is especially useful in the analysis of hydrolytic processes involving phosphorylated substrates.     

Direct spectrophotometric observation of an acyl-enzyme intermediate in elastase catalysis.

A technique is presented that is useful for selecting conditional-lethal mutants of mycoplasma cells and viruses. The method is based on growing mycoplasma on Millipore filters. Mutants can be isolated directly from filters seeded with mycoplasma. The filters can be transferred from condition to condition, acting as its own “master” and “replica” template. Virus mutants from the non-lytic Mycoplasma Group L1 and L2 viruses can also be picked from filters seeded with infected cells. This method is analogous to classical “replica plating” which is not a practical technique for mycoplasmas.     

Use of nile red for the rapid in situ quantitation of lipids on thin-layer chromatograms

We describe the use of the fluorescent dye nile red, 9-diethylamino-5H-benzo[alpha]phenoxazine-5-one, as a general-purpose reagent for the rapid detection and quantitation of a wide variety of lipids and other hydrophobic compounds separated by thin-layer chromatography. After samples are applied to silica gel plates and chromatographed, the plate is briefly dipped into a nile red solution (8 micrograms/ml of methanol-water 80:20, v/v). Background fluorescence of nile red dye adsorbed to the silica gel is then preferentially destroyed by dipping the plate in a dilute aqueous solution of bleach. After drying, lipid bands are visualized under ultraviolet light. Reflectance fluorometry (Ex: 580 nm; Em: 640 nm) is utilized for in situ quantitative analysis of the fluorescence of the lipids on the nile red-stained plate. Neutral lipids, phospholipids, sphingolipids, and fatty acids can be examined, although the nile red fluorescence intensity varies significantly among the lipid classes. Also, staining is stronger for unsaturated lipids than for saturated lipids. The lower detection limit of the assay is 25-100 ng for cholesterol, cholesteryl esters, triacylglycerols, and phospholipids.     

Direct spectrophotometric determination of mucin disulfide linkages

The method for determining S-S linkages in compounds of low molecular weight described by Zahler and Cleland is applied to purified porcine gastric mucin, a high molecular weight compound. The influence of the amount of starting material, atmospheric oxygen, and the reaction time were studied in order to establish the experimental conditions that give satisfactory results. The method described gave a value of 1.77 ± 0.07 mmoles SH per 100 g of mucin.     

The quantitation of rabies-specific antibodies. I. Modified counter immunoelectrophoresis. A rapid and sensitive method

Modified counter immunoelectrophoresis was standardized with respect to dilution of tissue culture antigen and indicator serum, the incubation time for neutralization and the effect of an electric current. The technique was found to be sensitive enough to detect a minimum level of antibodies (0.5 IU/ml) by using a 16 mA current per slide for 2 h, indicator serum of 15 IU/ml and the use of an antigen at a concentration of 1:35. Above all, the incubation period did not affect the neutralization of the virus. The test was also applied to the detection of rabies-specific antibody levels in 73 human sera. The test was found to be simple, quick and economical for titration of rabies antibodies.