Beckman Coulter COULTER LH 750 System, Справочник

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PN 4277248DD
1-5
USE AND FUNCTION
METHOD HISTORY
1
Conductivity Analysis
Cell walls act as conductors to high-frequency current. The current, while passing through
the cell walls and through each cell interior, detects differences in the insulating properties of
cell components. The current characterizes the nuclear and granular constituents and the
chemical composition of the cell interior.
20,21,22
Light Scatter Analysis
Coulter's experience in flow cytometry dates back decades to Fulwyler's pioneering use of
light scatter for cell analysis.
23
Loken et al. and Jovin et al. discuss the relationship of particle
size and refractivity to the angle of light scattered from a laser beam.
24,25
Reticulocyte (Retic) Analysis
Reticulocytes are immature, nonnucleated erythrocytes retaining a small network of
basophilic organelles, consisting of RNA and protoporphyrin. The enumeration of
reticulocytes provides a simple, effective means to determine red cell production and
regeneration.
26,27,28,29
The most common means of measuring reticulocytes is to use supravital dyes, such as New
Methylene Blue or Brilliant Cresyl Blue. These dyes precipitate and aggregate the basophilic
substances within the reticulocyte, resulting in a granular, staining pattern easily seen with
light microscopy.
30
Reticulocyte immaturity is related to cell volume and light scatter. Since more immature
reticulocytes are larger, contain more RNA and cause increased light scatter, the cell volume
and light scatter will increase with immaturity of the cell.
Figure 1.3 illustrates the IRF and MRV algorithms. This figure is a representation of the VCS
data that is shown on the two dimensional analyzer displays.
Figure 1.3 Illustration of the ten light scatter regions
The RET% is calculated as the ratio of reticulocytes to the total number of red cells. The
spectrum of light scatter intensity for the retic population is analyzed algorithmically. The
detected light scatter intensity of the retic population is divided into equal regions as shown
above. The IRF parameter is calculated as the ratio of the total number of retic events in the
outermost eight regions (3 to 10) to the total number of retics (regions 0 to 10 - region 0 is
not illustrated above). The MRV parameters is calcualted as the average volume of all
reticulocytes or the mean volume of all retic events.
1 1 0 2 3 4 5 6 7 8 9
M a t u r e
r e d c e l l s R e t i c u l o c y t e s
H i g h l i g h t s c a t t e r r e t i c u l o c y t e s
P l a t e l e t s / d e b r i s
M a t u r e
r e d c e l l s
R e t i c s
W h i t e
c e l l s
L i g h t s c a t t e r
Vo
lum
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