Lysing reagent, cartridge and automatic electronic cell counter for simultaneous enumeration of different types of white blood cells

• US 20060177347A1
• Filed: 06/11/2003
• Published: 08/10/2006
• Est. Priority Date: 06/11/2002
• Status: Active Grant

• Current Assignee: Koninklijke Philips Electronics N.V. (Koninklijke Philips N.V.)
• Original Assignee: Koninklijke Philips Electronics N.V. (Koninklijke Philips N.V.)
• Inventors: Hansen, Stine Hallberg, Larsen, Ulrik Darling, Ekberg, Bjorn
• Granted Patent: US 8,227,250 B2
• Time in Patent Office: Days
• US Class Current: 422/73
• CPC Class Codes: B01F 33/30 Micromixers
• CPC Class Codes: B01F 33/403 for mixing liquids B01F33/402, B01F33/405, B01F33/406 take precedence
• CPC Class Codes: B01L 2200/027 for microfluidic devices
• CPC Class Codes: B01L 2300/0645 Electrodes
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/0867 Multiple inlets and one sample wells, e.g. mixing, dilution
• CPC Class Codes: B01L 2300/087 Multiple sequential chambers
• CPC Class Codes: B01L 2400/0406 capillary forces
• CPC Class Codes: B01L 2400/0478 pistons
• CPC Class Codes: B01L 2400/0481 squeezing of channels or chambers
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: G01N 15/12 by observing changes in resistance or impedance across apertures when traversed by individual particles, e.g. by using the Coulter principle
• CPC Class Codes: G01N 15/1218 concerning the aperture
• CPC Class Codes: G01N 15/13 Details pertaining to apertures
• CPC Class Codes: G01N 2015/1024 Counting particles by non-optical means
• CPC Class Codes: G01N 2015/1029 Particle size
• CPC Class Codes: G01N 2015/1062 counting the particles by other than electro-optical means by electro-optical means G01N2015/1486
• CPC Class Codes: G01N 2015/1087 Particle size
• CPC Class Codes: G01N 2021/0321 One time use cells, e.g. integrally moulded
• CPC Class Codes: G01N 21/03 Cuvette constructions
• CPC Class Codes: G01N 21/35 : using infrared light G01N21/39 takes precedence
• CPC Class Codes: G01N 21/359 : using near infrared light
• CPC Class Codes: G01N 33/5094 : for blood cell populations red blood cells G01N33/80
• CPC Class Codes: G01N 33/80 : involving blood groups or blood types or red blood cells white blood cells G01N33/56972
• CPC Class Codes: Y10T 436/107497 : Preparation composition [e.g., lysing or precipitation, etc.]
• CPC Class Codes: Y10T 436/117497 : with a continuously flowing sample or carrier stream
• CPC Class Codes: Y10T 436/25625 : Dilution
• CPC Class Codes: Y10T 436/2575 : Volumetric liquid transfer

Abstract

The present invention relates to a lysing reagent for use in the simultaneous automatic electronic enumeration and volumetric discrimination of different types of blood cells, such as leukocytes, thrombocytes, etc, in blood. Further, the present invention relates to a Coulter (impedance) counting apparatus containing the lysing reagent, for example a Coulter counting apparatus with a disposable cartridge for characterizing cells suspended in a liquid, especially a self-contained disposable cartridge for single-use analysis, such as for single-use analysis of a small quantity of whole blood.

Claims

1. A lysing reagent for use in a blood cell counter for counting and discriminating a plurality of cell types in one counting operation, with a lysing capability sufficient for lysing of erythrocytes while maintaining counting ability of other cell types.

2. A lysing reagent mixture according to claim 1, wherein the lysing reagent contains a surfactant.

3. A lysing reagent mixture according to claim 1 or 2, wherein the surfactant comprises a saponin.

4. A lysing reagent according to claim 1, wherein the lysing reagent comprises a quaternary ammonium salt.

5. A lysing reagent according to any of claims 14, wherein the other cell types can be counted and discriminated In a Coulter counter.

6. A lysing reagent according to any of claims 1-5, wherein cells of the other cell types are reduced in size and the concentration is determined by counting a representative fraction of the respective cells.

7. A lysing reagent according to any of claims 1-6, wherein the other cell types include sub-populations of leukocytes, such as lymphocytes, monocytes and granulocytes, which are selectively reduced in size by the lysing reagent and can be counted in a cell counter.

8. An automatic electronic cell counter for counting and discriminating a plurality of blood cell types In one counting operation, having a chamber holding a lysing reagent according to any of the preceding claims.

9. A cartridge for characterizing blood cells in a blood sample, comprising a housing with

a first liquid storage chamber for holding a lysing reagent according to any of claims 1-7,

a first mixing chamber and a first collection chamber separated by a wall containing a first orifice for the passage of the cells between the first mixing chamber and the first collection chamber,

first cell characterization means for characterizing cells passing through the first orifice,

a bore in the outer surface of the housing for entrance of the blood sample, communicating with

a first sampling member positioned in the housing for sampling the blood sample and having a first cavity for receiving and holding the blood sample, the member being movably positioned in relation to the housing In such a way that, in a first position, the first cavity is in communication with the bore for entrance of the blood sample into the first cavity, and, in a second position, the first liquid storage chamber communicates through the first cavity with the first mixing chamber so that the blood sample can be flushed with discharged liquid from the first liquid storage chamber into the first mixing chamber.

10. A cartridge according to claim 9, further comprising

a second holding chamber and a second collection chamber separated by a second wall containing a second orifice for the passage of the cells between the second mixing chamber and the second collection chamber,

second cell characterization means for characterizing cells passing through the second orifice, and wherein

in the second position, the first cavity is in communication with the first mixing chamber for entrance of liquid from the first mixing chamber into the first cavity, and,

in a third position, the first cavity is in communication with the second mixing chamber for discharge of the liquid in the first cavity into the second mixing chamber.

11. A cartridge according to claim 9, further comprising

a second mixing chamber and a second collection chamber separated by a second wall containing a second orifice for the passage of the cells between the second mixing chamber and the second collection chamber,

second cell characterization means for characterizing cells passing through the second orifice, and

a second sampling member positioned in the housing for sampling a small and precise volume of liquid from the first mixing chamber and having a second cavity for receiving and holding the sampled liquid, the member being movably positioned In relation to the housing in such a way that, in a first position, the second cavity is in communication with the first mixing chamber for entrance of liquid from the first mixing chamber into the first cavity, and, In a second position, the second cavity is in communication with the second mixing chamber for discharge of the sampled liquid in the second cavity into the second mixing chamber.

12. A cartridge according to any of claims 9-11, further comprising a reagent chamber positioned adjacent to the first mixing chamber for holding a reagent to be entered into the first mixing chamber.

13. A cartridge according to claim 12, further comprising a breakable seal separating the reagent chamber from the first mixing chamber.

14. A cartridge according to any of claims 9-13, wherein a mixing member is positioned in at least one of the mixing chambers.

15. A cartridge according to any of claims 9-14, further comprising a sensor for characterization of the liquid.

16. A cartridge according to claim 15, wherein the sensor for characterization of the liquid is adapted for spectrophotometric characterization of the liquid.

17. A cartridge according to any of claims 9-16, wherein the orifice has a diameter in the range from 30 μ

m to 100 μ

m.

18. A cartridge according to claim 17, wherein the orifice has a diameter In the range from 35 μ

m to 50 μ

m.

19. A cartridge according to claim 17, wherein the orifice has a diameter in the range from 30 μ

m to 45 μ

m.

20. A cartridge according to claim 17, wherein the orifice has a diameter in the range from 35 μ

m to 40 μ

m.

21. A cartridge according to claim 17, wherein the orifice has a diameter substantially equal to 40 μ

m.