Mount Sinai School of Medicine

Miles Inc.

Bayer Corporation

Technicon Instruments Corporation

Grace E Martin

Leonard Ornstein

Gregory M Colella

Sophie S Fan

Gena Fischer

Daniel Ben-David

Albert Cupo

Young R Kim

Robert J Kanter

Hudson R Ansley

Methods for blood sample preparation using reagent compositions which include a zwitterionic surfactant for sphering of cells to eliminate orientational noise. When the reagent composition including a dye for staining a subset of cells is reacted with a blood sample, and the reaction mixture is passed through the sensing region of a flow cytometer, the light scattered and absorbed by each cell is measured, the stained cells can be distinguished from the unstained cells, and when the cells are reticulocytes and mature erythrocytes, respectively, the volume and hemoglobin concentration of each reticulocyte or erythrocyte, and the hemoglobin content, mean cell volume, mean corpuscular hemoglobin concentration, and mean cell hemoglobin of the reticulocytes or erythrocytes are calculated from the measured cell-by-cell volume and hemoglobin concentration.

Reagent compositions for use in sphering cells

A reagent composition which includes organic cationic dyes for staining the reticulocytes in a blood sample and buffer solutions for maintaining a pH of about 6 to about 9 is provided. The dyes may be the blue absorption dyes Oxazine 750 or New Methylene Blue. A zwitterionic surfactant is included in the reagent composition for isovolumetric sphering of the reticulocytes and erythrocytes. The reagent composition and whole blood sample mixture are passed through the sensing region of a flow cytometer. The light scattered and absorbed by each cell is measured; the erythrocytes can be distinguished from reticulocytes and the volume, hemoglobin concentration and the hemoglobin content of each reticulocyte or erythrocyte, and the mean cell volume, mean corpuscular hemoglobin concentration, and mean cell hemoglobin of the reticulocytes or erythrocytes are calculated from the measured cell-by-cell volume and hemoglobin concentration.

Reagent compositions for use in the identification and characterization

Methods for characterizing and distinguishing reticulocytes and mature red blood cells use reagent compositions which include organic cationic dyes for staining the reticulocytes in the blood sample and buffer solutions for maintaining a pH of about 6 to about 9. The dyes may be the blue absorption dyes Oxazine 750 or New Methylene Blue. When a zwitterionic surfactant is included in the reagent composition for isovolumetric sphering of the reticulocytes and erythrocytes, and the reagent composition and whole blood sample mixture is passed through the sensing region of a flow cytometer, the light scattered and absorbed by each cell is measured, the erythrocytes can be distinguished from reticulocytes and the volume, hemoglobin concentration and the hemoglobin content of each reticulocyte or erythrocyte, and the mean cell volume, mean corpuscular hemoglobin concentration, and mean cell hemoglobin of the reticulocytes or erythrocytes are calculated from the measured cell-by-cell volume and hemoglobin concentration. One method also includes adjusting the measured absorption signals for pseudo-absorption.

Methods for the identification and characterization of reticulocytes in

Methods for blood sample preparation using reagent compositions which include a zwitterionic surfactant for sphering of cells to eliminate orientational noise. When the reagent composition including a for staining a subset of cells is reacted with an anticoagulated blood sample, and the reaction mixture is passed through the sensing region of a flow cytometer, the light scattered and absorbed by each cell is measured, the stained cells can be distinguished from the unstained cells, and when the cells are reticulocytes and mature erythrocytes, respectively, the volume and hemoglobin concentration of each reticulocyte or erythrocyte, and the hemoglobin content, mean cell volume, mean corpuscular hemoglobin concentration, and mean cell hemoglobin of the reticulocytes or erythrocytes are calculated from the measured cell-by-cell volume and hemoglobin concentration.

Reagent compositions and their use in sphering cells

A flow cytometry method for reproducibly detecting and counting a lymphocyte population of interest in a leukocyte suspension or whole blood sample in which the red cells are subsequently lysed. The suspension (or sample) is combined with a reagent comprising a primary antibody, either native, carrying an attached enzyme or biotin or other label, and a fixative reagent, in either order. Where the enzyme is not attached, an enzyme is coupled specifically to the primary antibody. The fixed suspension is reacted with a color-producing enzyme-cytochemical reagent. The suspension, now including stained and unstained fixed cells, is passed through a flow cytometer and the cells are characterized and counted on the basis of their light-scattering and light-absorbing properties.

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Mount Kisco, NY, United States of America

Grace E Martin