Method and apparatus for analyzing individual cells or particulates using fluorescent quenching and/or bleaching
A method for analyzing a blood sample is provided that includes the steps of: providing a blood sample having one or more of each first and second constituents; admixing a colorant with the sample, which colorant is operative to cause the first constituents and second constituents to fluoresce and absorb light; illuminating at least a portion of the sample; e) imaging a portion of the sample; determining a fluorescence value for each the first constituents and second constituents; determining an optical density value for each of the first constituents and second constituents; and identifying the first constituents and the second constituents using the determined fluorescence and optical density values.
1. A method for analyzing at least one constituent of a biologic fluid sample quiescently residing within an analysis chamber defined by a first panel and a second panel, and which sample is mixed with at least one colorant that is operable to reside within the constituent and emit fluorescent light upon exposure to predetermined first wavelengths of light, the method comprising the steps of:
illuminating at least a portion of the sample containing the constituent at one or more first wavelengths in a constant manner for a period of time;
imaging the at least a portion of the sample at discrete points in time during the period, including producing image signals at the discrete points in time, which signals are indicative of fluorescent emissions from colorant residing within the constituent;
determining one or more fluorescent emission values associated with the colorant residing within constituent using the image signals from the discrete points in time, and a rate of change for the fluorescent emission values between a plurality of the discrete points in time; and
analyzing the constituent using the determined rate of change of the fluorescent emission values for the constituent.
2. The method of claim 1 , wherein the analyzing step includes determining a characteristic of the constituent.
3. The method of claim 2 , wherein the characteristic is an age of the constituent.
4. The method of claim 1 , wherein the at least one constituent includes a first constituent and a second constituent;
wherein the step of determining one or more fluorescent emission values includes determining the one or more fluorescent emission values from the colorant residing within the first constituent and from the second constituent using the image signals from the discrete points in time, and a rate of change for the fluorescent emission values between a plurality of the discrete points in time for each of the first constituent and the second constituent; and
analyzing the first constituent and the second constituent using the determined rate of change of the fluorescent emission values for the respective constituent.
5. The method of claim 4 , wherein the first constituent and the second constituent are a same type of constituent.
6. The method of claim 5 , wherein the analyzing step includes determining a characteristic of the first constituent and the second constituent.
7. The method of claim 6 , wherein the characteristic is an age of the respective constituent.
8. The method of claim 4 , wherein the biologic fluid sample is substantially undiluted whole blood.
9. The method of claim 8 , wherein the first and second constituents are selected from the group consisting of white blood cells and particulates.
10. The method of claim 9 , wherein the first and second constituents are each types of white blood cells.
11. The method of claim 10 , wherein the first constituents and the second constituents are selected from the group consisting of eosinophils, neutrophils, and basophils.
12. The method of claim 9 , wherein the first and second constituents are each types of particulates.
13. The method of claim 12 , wherein the first and second particulates are selected from a group consisting of platelets and hematoparasites.
14. The method of claim 1 , wherein the one or more one or more values representative of fluorescent emission from the constituent are determined on a per pixel basis.
15. The method of claim 1 , wherein the steps of determining one or more fluorescent emission values and analyzing the constituent are performed using one or more processors adapted to process the image signals.
16. The method of claim 1 , wherein the step of determining one or more fluorescent emission values associated with the colorant includes determining fluorescent emission values at different wavelengths.
17. An apparatus for analyzing a blood sample, comprising:
a chamber adapted to quiescently hold the sample;
an illuminator operable to illuminate at least a portion of the sample containing a constituent, along one or more fluorescent excitation wavelengths in a constant manner for a period of time;
an image dissector operable to image the at least a portion of the sample at discrete points in time during the period, and produce image signals representative of fluorescent emissions from the constituent at the discrete points in time; and
an analysis device adapted to determine one or more fluorescent emission values for the constituent quiescently disposed within the sample using the image signals at the discrete points in time, and a rate of change for the fluorescent emission values between the discrete points in time, and adapted analyze the constituent using the determined rate of change of the fluorescent emission values for the constituent.
18. The apparatus of claim 17 , wherein the constituent is selected from the group consisting of white blood cells and particulates.
19. The apparatus of claim 17 , wherein the analysis device is operable to determine a characteristic of the constituent, using the image signals at the discrete points in time, and the rate of change for the fluorescent emission values between the discrete points in time.
20. The apparatus of claim 19 , wherein the characteristic is an age of the constituent.