Effects of phosphatidylinositol diphosphate on phospholipid asymmetry in the human erythrocyte membrane.

While phospholipid asymmetry has been well characterized in red blood cells (RBCs), controversy exists as to what role PIP2 plays in cation-induced phosphatidylserine (PS) exposure. We report that PIP2 can redistribute intracellular cations and thereby lead to a loss of phospholipid asymmetry. Flow cytometry was employed to monitor intracellular cation levels by using the fluorophore Fluo-3 and exposure of PS on the outer surface of the RBC bilayer by using fluorescently labeled annexin V.

Colorimetric determination of DNase I activity with a DNA-methyl green substrate.

A simple, high throughput, and precise assay was developed for quantification of deoxyribonuclease I (DNase; IUB 3.1.21.

Capillary electrophoresis of abnormal hemoglobins associated with alpha-thalassemias.

Capillary electrophoresis was evaluated for separation of hemoglobin species associated with alpha-thalassemias, and for identification of hemoglobin variants commonly found in the same human populations. Separation of hemoglobins was achieved using capillary isoelectric focusing with chemical mobilization; visible-wavelength absorbance detection was used to identify hemoglobins against a background of nonheme-containing proteins. This technique could easily differentiate hemoglobins Bart's and H (associated with alpha-thalassemias) from hemoglobin variants.

Lung surfactant proteins, SP-B and SP-C, alter the thermodynamic properties of phospholipid membranes: a differential calorimetry study.

The ability of the low molecular weight lung surfactant-associated proteins, SP-B and SP-C, to alter the thermotropic properties of synthetic multilamellar vesicles was tested using differential scanning calorimetry (DSC). The presence of either SP-B or SP-C in dipalmitoylphosphatidylcholine (DPPC) or dipalmitoylphosphatidylglycerol (DPPG) multilamellar vesicles broadened the DSC thermogram and reduced the enthalpy of transition in a concentration-dependent manner. With both proteins, the temperature at which the peak of the phase transition (Tm) was detected was shifted to a higher value.

Improved isolation of normal human reticulocytes via exploitation of chloride-dependent potassium transport.

Studies on normal human reticulocytes have been limited by a lack of methods for effective reticulocyte enrichment. This study shows a convenient new approach for selective enrichment of reticulocytes from normal blood samples. We have developed a modified arabinogalactan density gradient that contains high potassium levels, approximating the internal cation composition of red blood cells (RBC).