Abstract: A single laser flow cytometric method for the enumeration of micronuclei in erythrocyte populations, wherein a sample of peripheral blood or bone marrow is obtained and the cell populations in the sample are fixed. Reticulocytes in the fixed samples are treated simultaneously with RNAse and with a fluorescent labeled antibody having binding specificity for a surface marker for erythroblasts/reticulocytes. The erythrocyte populations are then stained with a nucleic acid stain which stains DNA representing micronuclei, if present. The stained and/or labelled erythrocyte populations are then exposed to a laser beam of appropriate excitation wavelength for both the nucleic acid staining dye and the fluorescent label to produce fluorescent emission. The fluorescent emission and light scatter produced by the erythrocyte populations are detected by the flow cytometer from which is calculated the number of specific erythrocyte populations in said sample.
Type:
Grant
Filed:
September 6, 1996
Date of Patent:
January 12, 1999
Assignee:
Litron Laboratories
Inventors:
Stephen Dertinger, Dorothea Torous, Kenneth Tometsko
Abstract: Disclosed are a composition for the lyophilization of mammalian red blood cells comprising a hydrophilic polymer, a carbohydrate, and an organic solvent; and a method of using the composition to lyophilize red blood cells comprising mixing red blood cells with the composition, freezing the mixture, and drying the mixture by removing water by sublimation. Also disclosed are red blood cells lyophilized according to this method for lyophilization, and a method for reconstituting the lyophilized red blood cells. In particular, the composition used to lyophilize the red blood cells comprises a mixture of a hydrophilic polymer ranging from 1,450-20,000 Daltons at 5-50% w/v, a mono- or disaccharide or a mixture thereof from 0.01-0.2M and an organic solvent such as a primary alcohol, a secondary alcohol, dimethyl sulfoxide or combinations thereof at 0.5-20% v/v.
Type:
Grant
Filed:
June 19, 1996
Date of Patent:
May 12, 1998
Assignee:
Litron Laboratories
Inventors:
Andrew M. Tometsko, deceased, Stephen Dertinger, Dorothea Torous, Kenneth Tometsko
Abstract: Genotoxic chemicals are an existing wide-spread health hazard to the human population. Advances in genetic toxicology testing have made it possible to assay potential mutagens, carcinogens, teratogens and clastogens in the environment. The mouse micronucleus assay provides an example of an excellent test for genetic damage to cells. When chromosome breaks occur in the blood stem cell population, the damaged piece of chromosome remains behind as a micronucleus in the normally DNA deficient red blood cells. However, currently available manual micronucleus assays are costly, time consuming, and labor intensive. In addition, the statistics are often marginal since the number of micronucleii (MNs) in 1000 polychromatic cells are scored manually, yielding limited amounts of data. This invention discloses the means for assaying the change in micronucleated cells by high speed flow cytometry.