Identification of blast cells in a leukocyte cell preparation
AssigneeCoulter International Corp.
InventorsNazle Kuylen, Harold R. Crews, Gerald E. Jaffe, Maria Barcelon, Francis J. Lucas, Nancy M. Garcia
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Abstract
The necessity to detect blast cells in a patient sample to determine a treatment regime for leukemia has long been desired. The present invention relates to a method and assay reagents for identification of blast cells in a fluid sample. The method further enables the determination of the lineage of the blast cells. The new method provides a determination of blast cells in less time than prior art methods.
Claims34
A method for identification of blast cells in a fluid sample comprising:
- a) taking a body fluid sample from an individual, said sample containing metabolically active cells,
- b) preparing said fluid sample to form at least one test sample, said test sample containing metabolically active cells,
- c) adding an assay reagent to each prepared test sample to form a test cell mixture, said assay reagent containing a fluorogenic indicator group being selected for its ability to have a non-fluorescent first state before said assay reagent reacts with an enzyme within said cells, and a fluorescent second state excitable at a wavelength above 450 nm after said assay reagent reacts with said enzyme within said cell, said assay reagent having a fluorescence less than the auto-fluorescence of a metabolically active cell before reacting with said enzyme within said cell,
- d) illuminating said test cell mixture with a beam of light which causes said light to be scattered and exciting said test cell mixture with a fluorescent wavelength above 450 nm which causes said test cell mixture to have a fluorescent second state,
- e) detecting fluorescence and light scatter of the cells in said test cell mixture, and
- f) correlating said fluorescence and said light scattering of said cells to identify blast cells contained is said test cell mixture.
An assay reagent for identification of blast cells in a fluid sample by measurements of cellular enzyme activity comprising:
- a) an assay compound selected from 3', 6' fluorescein dichloroacetate and 3', 6' fluorescein dibutyrate;
- and
- b) a physiological buffer, wherein said assay reagent has a fluorescence less than autofluorescence of a metabolically active cell before reacting with an enzyme within a cell and a pH from 4 to 7.5.
Dependencies are inferred from the claim text.