The receptor for transferrin on murine myeloma cells: one-step purification based on its physiology, and partial amino acid sequence

The receptor for transferrin is one of the major surface proteins of proliferating lymphocytes and other cells. It binds ferrotransferrin from serum and endocytoses it into an acidic nonlysosomal intracellular compartment where iron is released, but in which apotransferrin remains tightly bound to its receptor. Recycling of the apotransferrin-receptor complex to the cell surface is associated with a return to neutral pH and concomitant loss of affinity of apotransferrin for its receptor. Apotransferrin is then free to leave the cell and initiate a new cycle. We have exploited this cycle in a novel method for the purification of the receptor for transferrin. Murine myeloma cells were lysed in nonionic detergent, and the lysate passed over a column of ferrotransferrin-agarose at pH 7.4. After washing with sodium acetate at pH 5.0, iron was removed with sodium citrate pH 5.0 and desferrioxamine. Upon returning the pH to neutrality, the receptor was eluted and found to be homogeneous by SDS-polyacrylamide gel electrophoresis under both reducing and nonreducing conditions. The degree of purification was estimated to be at least 3,000-fold, and the calculated yield was 10 to 20%. The purified receptor was capable of binding to transferrin. The receptor was digested with trypsin, and the resulting peptides were separated by reversed-phase high performance liquid chromatography in NH4HCO3. Selected peptides were rechromatographed in 0.1% trifluoroacetic acid, and their amino acid sequences were determined.     

Population structure,dispersal and colonization history of the garden bumblebee <Emphasis Type="Italic">Bombus hortorum</Emphasis> in the Western Isles of Scotland

New methods of analysing genetic data provide powerful tools for quantifying dispersal patterns and reconstructing population histories. Here we examine the population structure of the bumblebee Bombus hortorum in a model island system, the Western Isles of Scotland, using microsatellite markers. Following declines in other species, B. hortorum is the only remaining long-tongued bumblebee species found in much of Europe, and thus it is of particular ecological importance. Our data suggest that populations of B. hortorum in western Scotland exist as distinct genetic clusters occupying groups of nearby islands. Population structuring was higher than for other bumblebee species which have previously been studied in this same island group (F_st = 0.16). Populations showed significant isolation by distance. This relationship was greatly improved by using circuit theory to allow dispersal rates to differ over different landscape features; as we would predict, sea appears to provide far higher resistance to dispersal than land. Incorporating bathymetry data improved the fit of the model further; populations separated by shallow seas are more genetically similar than those separated by deeper seas. We argue that this probably reflects events following the last ice age when the islands were first colonized by this bee species (8,500–5,000 ybp), when the sea levels were lower and islands separated by shallow channels would have been joined. In the absence of significant gene flow these genetic clusters appear to have since diverged over the following 5,000 years and arguably may now represent locally adapted races, some occurring on single islands.     

Development of a diagnostic test system for detecting soluble staphylococcal antigens by an immunoenzyme method

ELISA is used for detecting the soluble staphylococcal antigen in patients with purulent septic infections. The optimum conditions for the assay have been established: the dose of staphylococcal gamma globulin for plate sensitization should be 5.0-10.0 micrograms/ml, the pH of the buffer solution 9.6-10.0, the time and temperature of incubation 18-20 hours at 4 degrees C or 5 hours at 37 degrees C. The possibility of using plates manufactured in the USSR has been shown. The sensitivity of the above diagnostic test system is 0.005 microgram/ml.     

Simple reversed-phase high-performance liquid chromatography quantitation of ganciclovir in human serum and urine

A fast, simple, and cost-effective HPLC method for the quantitation of the antiviral drug ganciclovir is described. The serum samples are extracted with perchloric acid and neutralized with potassium phosphate buffer, and urine samples are diluted with distilled water. A reversed-phase column with isocratic elution by 15 mM potassium phosphate buffer (pH 2.5) containing 0.25% acetonitrile is used to separate ganciclovir; quantitation is by UV absorbance at 254 nm. Total turnaround time is 22 min; more than 3000 samples can be run on a single column without loss of peak quality. The limit of quantitation is 0.05 μg/ml. Recoveries varied from 91 to 10% with coefficients of variation ranging from 0.387 to 7.95%.