COULTER HmX Hematology Analyzer, Reference Manual

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PN 4237523A 3-2
OPERATION PRINCIPLES
GENERAL PRINCIPLES
Figure 3.2 Flow Cell
Effect of Reagents
The conductive diluent must affect cells minimally, if at all.
Both lytic reagents must destroy erythrocytes without significantly affecting leukocytes. They
must work rapidly to satisfy the speed with which the system works.
The leukocyte preservative must:
r
Provide clear separation of the white blood cell populations
r
Preserve leukocytes in their near-native state for accurate cytometric measurement.
Retic Analysis
The Coulter Reticulocyte Method is a two-step sample preparation, followed by analysis in
the HmX Hematology Analyzer Retic Mode using volume conductivity and scatter (VCS)
technology. First, a supravital dye, New Methylene Blue, in a special solution (Reagent A), is
incubated with whole blood samples. The dye precipitates the basophilic RNA network found
in reticulocytes. Then, when added to the samples, a hypotonic clearing reagent (Reagent B)
clears hemoglobin and unbound stain from the cells. After the treatment, and if viewed in a
wet prep microscopically, mature erythrocytes appear as clear, slightly spherical cells.
Reticulocytes have the same characteristics but with darkly-stained reticulum.
Sheath fluid
upper circuit
Sheath fluid
lower circuit
Sample
Waste
Aperture