Hematology

Analyzer
Automated Cell Counting
Instrumentation

Presented
Presented to:
to:
Mam
Mam Raheela
Raheela

Presented
Presented by:
by:
NAYAB
NAYAB ARSHAD
ARSHAD
2011-Dvme-001`
2011-Dvme-001`
Definition:
Hematology analyzers are computerized,
highly specialized and automated machines
that count the number of different kinds of
white and red blood cells in a blood sample.
History:
The original
hematology analyzers
first appeared in the
1950s, but the truly
functional and
automated versions
of these machines did
not become available
for two more
decades. Before this
time, cell counts were
performed manually.
Disadvantages of manual
cell counting
Cell identification errors in manual counting:
Mostly associated with distinguishing
lymphocytes from monocytes
Bands from segmented forms and abnormal
cells (variant lymphocytes from blasts)
Lymphocytes overestimated
Monocytes underestimated
Slide cell distribution error
Increased cell concentration along edges
Advantages
They have a high level of
precision for cell counting and
cell sizing greatly superior to that
of the manual technology
The results are generally
accurate.
No slide distribution error.
Eliminate statistical
Continu variations associated with
manual count based on high
e number of cells counted.
Many parameters are available.
All automated cell counters are
screening devices.
(abnormalities)
Advantages..
Red blood cell (RBC) counts
Hemoglobin in RBCs,
Hemocrit levels,
Platelet counts,
Corpuscular data,
Counts of five different types of
white blood cells.
Most current Some newer machines also
hematology measure numbers of two
analyzers specialized cell types
provide immature white cells and
nucleated red blood cells that
are necessary to confirm
specific diagnoses
Results:
The results they provide are collectively
known as complete blood counts (CBCs) .
complete complete blood with differentiation
of cells (CBCs with diff).
Some cell counters can process 120-150
samples per hour .
Hematology
analyzer has
revolutionised
Types :
Automated techniques of blood counting:

I. Semi-automated instruments
Require some steps, as dilution of blood samples.
Often measure only a small number of variables.
II. Fully automated instruments
Require only that an appropriate blood sample is
presented to the instrument.
They can measure 8-20 variables including
some new parameters which do not have any
equivalent in manual methods.
Working
Hematology analyzers count cells by one
of five general cell techniques:
Flow cytometry
The use of fluorescent dyes
The electrical impedance method
The light scatter method
Radiofrequency
1:Flow cytometry/Cell
sorting
o Flow cytometry is based on ejecting cells
from a nozzle at high speed in a fluid.
o Each cell passes through several laser beams
so that different optical properties can be
measured.
Continue
2:Use of flourescent
dyes
Biochemical or antigenic properties are
usually classified using fluorescent dyes.
Continue.
Steps :
1. Labeling with flourescent dye.
2. A laser excites these fluorescent molecules.
3. They emit light at various wavelengths.
4. Amount of fluorescence can give an
indication as to what percentage of various
cell types are present in the sample.
3:Electrical
impedance method
Works on coulter principle:
A stream of cells in suspension passes through a
small aperture across which an electrical current
is applied. Each cell that passes alters the
electrical impedance and can thus be counted and
sized.
Continue
Particles such as blood cells are nonconductive
BUT
are suspended in an electrically conductive
diluent.

As a dilute suspension of cells is drawn through

the aperture, the passage of each individual cell
momentarily increases the impedance
(resistance) of the electrical path between two
electrodes that are located on each si.de of the
aperture
Continue
A blood cell's size ,
surface charge ,
concentration of the cells ,
shape of cells can be determined
4:Optical scatter
method
The application of light scatter means that as a
single cell passes across a laser light beam
1. diffraction(bending around corners),
2. refraction (bending due to change in speed) and
3. reflection (light rays turned back by
obstruction)
.Light scatter correlates to cell volume/size
Side angle/orthogonal light scatter correlates
to degree of internal complexity (granules and
nucleus)
Continue
Continue
The patterns of scatter are measured at various
angles.

Multi angle polarized scatter separation

(M.A.P.S.S)

0 :
indicator of cell size
10 :
indicator of cell structure and complexity
90 polarized:
indicates nuclear lobularity
90 depolarized:
differentiates eosinophils
Continue
Scattered light provides information about

cell structure,
shape,
Reflectivity
These characteristics can be used to
differentiate the various types of blood
cells and to produce scatter plots with a
five-part differential.
5:Radiofrequency
Examples
o An automated retic
count
Fluoro-chromes combine with the RNA of the
reticulocytes.
Fluorescent cells can then be enumerated
using a flowcytometer.
An automated retic counter also permits the
assessment of retic maturity since the more
immature reticulocytes have more RNA
o Coulter Instrumentation
Coulter hematology analyzer
 Direct Measurement:
RBC DC detection (Impedance),
hydrodynamic focusing
WBC DC detection (Impedance),
hydrodynamic focusing
Platelets DC detection
(Impedance), hydrodynamic
Principles focusing
(2-30 fl)

of Hgb SLS-Hb (555 nm)

(oxyhemoglobin + sodium lauryl
sulfate)
Measurem HCT cumulative pulse height
detection (see oscilloscope)

ent Indirect Measurement:

MCV, MCHC, and MCH (calculation)
WBC differential
DC/RF detection
employs differential lysis
Reticulocyte
Fluorescent detection
Symax hematology analyzer
Automated and portable hematology
analyzers
Drawbacks
Some problems which could be faced:
Two cells passing through the orifice at the
same time, counted as one cell.
RBC agglutination(clump of cells) Counting
bubbles
or other particles as cells.
Questions????
Thank
you