US4751179A
Expired / Lapsed
US
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Method and reagents for differential determination of four populations of leukocytes in blood
Assignee: Beckman Coulter Inc
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Filing: 1984-05-31
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Granted: 1988-06-14
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Expires: 2004-05-31
Abstract
Abstract This invention describes a reagent system, including saponin in a lysing reagent and a rapidly active cross-linking agent such as glutaraldehyde as a fixing reagent, which reproducably affects whole blood to cause the red blood cells to stromatolyze and modifies the leukocytes to generate data to define four distinct clusters for detection and classification by flow analysis instrumentation. The clusters represent the four major leukocyte types found in blood: lymphocytes, monocytes, neutrophils and eosinophils, thus providing a method of leukocyte differential analysis. The parameters used for the leukocyte classification include combinations of two or more of DC (Coulter) volume, high frequency (RF) size, opacity (RF size/DC volume), light scatter at various angular ranges, and fluorescence at various wavelengths of illumination.
Inventors
- 👤 Stephen L. Ledis
- 👤 Harold R. Crews
- 👤 Ted Sena
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-26 20:32
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/5005; G01N33/5094; Y; Y10; Y10T; Y10T436/00; Y10T436/10; Y10T436/101666; Y10T436/107497
Claims Analysis
Total Claims: 9
Claim 1
Independent Claim
148 words
A method for stromatolysing the red blood cells in a sample of whole blood and for classification of leukocytes into four categories, one of said categories being eosinophils, which comprises the steps of: I. treating a sample of whole blood with aqueous solutions of the following components: (A) a lysing reagent comprising (1) a single lytic diluent containing saponin, or (2) two solutions to be added separately to said sample of whole blood, consisting of (a) a prediluent and (b) a lytic reagent containing saponin; and then with (B) a fixing reagent containing a cross-linking compound; each of said components (A) and (B) being maintained at a predetermined concentration, pH and osmolality; II. measuring by instrumentation at least two of the primary parameters of DC volume, RF size, fluorescence, and light scatter, to produce cytograms; and III. observing four populations of leukocytes, one of said categories being eosinophils.
Claim 2
Independent Claim
148 words
A method for stromatolysing the red blood cells in a sample of whole blood and for classification of leukocytes into four categories, one of said categories being eosinophils, which comprises the steps of: I. treating a sample of whole blood with aqueous solutions of the following components: (A) a lysing reagent comprising (1) a single lytic diluent containing saponin, or (2) two solutions to be added separately to said sample of whole blood, consisting of (a) a prediluent and (b) a lytic reagent containing saponin; and then with (B) a fixing reagent containing a cross-linking compound; each of said components (A) and (B) being maintained at a predetermined concentration, pH and osmolality; II. measuring by instrumentation at least two of the primary parameters of DC volume, RF size, fluorescence, and light scatter, to produce cytograms; and III. observing four populations of leukocytes, one of said categories being eosinophils.
Claim 3
Dependent Claim
19 words
The method of claim 1 which further includes correlating the measured parameters of RF size/DC volume vs DC volume.
Claim 4
Dependent Claim
18 words
The method of claim 1 which further includes correlating the measured parameters of light scatter vs DC volume.
Claim 5
Dependent Claim
19 words
The method of claim 1 which further includes correlating the measured parameters of light scatter/DC volume vs DC volume.
Claim 6
Dependent Claim
17 words
The method of claim 1 which further includes correlating the measured parameters of fluorescence vs light scatter.
Claim 7
Dependent Claim
19 words
The method of claim 1 which further includes correlating of the measured parameters of fluoresence/light scatter vs light scatter.
Claim 8
Dependent Claim
16 words
The method of claim 1 wherein said categories of leukocytes are: lymphocytes, monocytes, neutrophils and eosinophils.
Claim 9
Dependent Claim
44 words
The method of claim 1 wherein the blood sample which has been treated as in step I is heated to an elevated temperature of about 60° to 75° C. for about 10 to 30 seconds prior to measurement by instrumentation as in step II.
References Cited (Backward Citations)
US2656508A
Cited
US3259842A
Cited
US3446751A
Cited
US3502974A
Cited
US3741875A
Cited
US3836849A
Cited
US3710933A
Cited
US4146604A
Cited
US3884579A
Cited
US3883247A
Cited
US3873467A
Cited
US4198206A
Cited
US4099917A
Cited
US4213876A
Cited
US4286963A
Cited
US4521518A
Cited
US4500509A
Cited
US4492752A
Cited
US4506018A
Cited
US4485175A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1991-06-14 | 1991-12-14 | 1992-06-14 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1995-06-14 | 1995-12-14 | 1996-06-14 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1999-06-14 | 1999-12-14 | 2000-06-14 | $7700.00 | Window Closed / Lapsed |