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ROCHESTER, NY, United States

A method for the enumeration of micronucleated erythrocyte populations while distinguishing platelet and platelet-associated aggregates involves the use of a first fluorescent labeled antibody having binding specificity for a surface marker for reticulocytes, a second fluorescent labeled antibody having binding specificity for a surface marker for platelets, and a nucleic acid staining dye that stains DNA (micronuclei) in erythrocyte populations. Because the fluorescent emission spectra of the first and second fluorescent labeled antibodies do not substantially overlap with one another or with the emission spectra of the nucleic acid staining dye, upon excitation of the labels and dye it is possible to detect the fluorescent emission and light scatter produced by the erythrocyte populations and platelets, and count the number of cells from one or more erythrocyte populations in said sample. In particular, the use of the second antibody prevents interference by platelet-associated aggregates in the scoring procedures.


The present invention relates a method for assessing in vivo hematotoxicity. The method utilizes differential staining of nucleated and non-nucleated blood cells, and also differential labeling of cells with functional versus dysfunctional mitochondrial membrane potential. Quantitative analyses can be conducted on stained whole blood specimens, and is based on blood cells fluorescent emission and light scatter properties following exposure to an excitatory light source. The ratio of certain cell populations can be readily measured. Furthermore, it is also possible to express cell population values in terms of number per unit volume. This invention can be used to evaluate the hematotoxicity of drugs, chemicals, radiation, and other exogenous agents, or the effects that a suspected protective agent may have on induced hematotoxicity. Furthermore, the matrix of measurements provided by this invention is useful in estimating radiation dose, i.e., retrospectively. Kits for practicing the invention are also disclosed.


Patent
Litron Laboratories, Ltd. | Date: 2010-10-06

The present invention relates a method for the enumeration of in vivo gene mutation. The method utilizes differential staining of GPI-anchor deficient erythrocyte populations to distinguish between wild-type and pig-a gene mutants. Quantitative analyses can be conducted on erythrocytes and/or reticulocytes, and is based upon fluorescent emission and light scatter following exposure to an excitatory light source. Counting of mutant erythrocytes or reticulocytes relative to the number of total erythrocytes or reticulocytes can be used to assess the DNA-damaging potential of an exogenous chemical agent, the DNA-damaging potential of an exogenous physical agent, the effects of an exogenous agent which can modify endogenously-induced DNA damage, and the effects of an exogenous agent which can modify exogenously-induced DNA damage. Kits for practicing the invention are also disclosed.


The present invention relates a method for the enumeration of mammalian cell micronuclei, while distinguishing micronuclei from the chromatin of dead and dying cells. The method utilizes differential staining of chromatin from dead and dying cells, to distinguish the chromatin from micronuclei and nuclei that can be detected based upon fluorescent emission and light scatter following exposure to an excitatory light source. Counting of micronuclei events relative to the number of nuclei can be used to assess the DNA-damaging potential of a chemical agent, the DNA-damaging potential of a physical agent, the effects of an agent which can modify endogenously-induced DNA damage, and the effects of an agent which can modify exogenously-induced DNA damage. Kits for practicing the invention are also disclosed.


A method for the enumeration of micronucleated erythrocyte populations while distinguishing platelet and platelet-associated aggregates involves the use of a first fluorescent labeled antibody having binding specificity for a surface marker for reticulocytes, a second fluorescent labeled antibody having binding specificity for a surface marker for platelets, and a nucleic acid staining dye that stains DNA (micronuclei) in erythrocyte populations. Because the fluorescent emission spectra of the first and second fluorescent labeled antibodies do not substantially overlap with one another or with the emission spectra of the nucleic acid staining dye, upon excitation of the labels and dye it is possible to detect the fluorescent emission and light scatter produced by the erythrocyte populations and platelets, and count the number of cells from one or more erythrocyte populations in said sample. In particular, the use of the second antibody prevents interference by platelet-associated aggregates in the scoring procedures.

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