Erythrocytes aka RBC’s
Laboratory Procedures
What is Blood?
Whole Blood = fluid and cells
Fluid Component = PLASMA
Cellular Component
• Erythrocytes (RBC)
• Leukocytes (WBC)
• Thrombocytes (Platelets)
How Are These Cells
Produced?
• Hematopoesis : the production of blood
cells and platelets.
• Erythropoesis : the production of
erythrocytes (RBC)
In juveniles, blood produced in
liver, spleen, thymus and bone
marrow.
In adults, primary site of
erythropoesis is in the bone marrow.
During times of hematopoetic stress,
the slpeen and liver may produce RBC
Leukopoiesis : the
production of leukocytes
or WBC
Thrombopoiesis : the
production of
thrombocytes or platelets
All of these cells come from
ONE CELL!!
Proteins called Cytokines
are responsible in
determining the fate of all
stem cells.
These cytokines determine in the cell will
be either :
Myeloid :
Erythroblasts – Erythrocytes
(Erythropoietin)
Megakaryocytes –Platelets
Myeloblasts –Leukocytes / Monocytes
Cellular
Maturation
Diagram
Blood Composition
• Separates into three components:
• Red Blood Cells (RBC’s)
• White Blood Cells and platelets (buffy
coat)
• Plasma
• Bottom 1/3 to ½ of tube contains the
heaviest of cellular material (the RBC’s).
Hematocrit=PCV (Packed Cell
Volume)
• To determine hematocrit, whole blood is
centrifuged to pellet the red blood cells.
• Plasma remains on the top of the red cells.
• The fraction of blood that is packed is the
hematocrit and is read as a percentage.
Hemoglobin (Hgb)
• Normal values are usually 1/3 of the hematocrit.
• Each hemoglobin molecule has 4 heme units
attached to globulins.
• Abnormal heme groups, cannot carry oxygen.
• Carboxyhemaglobin- Hgb has a higher affinity for
CO than O2.
• Bright red blood
• Methemoglobin- The Fe molecule is oxidized to
Fe+3.
• Blood becomes brown.
• Tylenol toxicity in cats.
Hemoglobin Molecule
Red Blood Cells
• Function:
• Carry oxygen to the tissues
• Oxygen must be carried at enough
pressure to permit rapid diffusion of
oxygen.
• The RBC is a vehicle for hemoglobin
which is the carrier molecule for
oxygen.
• The sigmoid shape of curve is a result
of the cooperative binding from the 4
hemoglobin molecules.
Erythrocytes
The morphological features
of mature red blood cells of
dogs, cats, horses, and
ruminants are generally very
similar in that they all lack
nuclei, stain reddish to
reddish-orange.
Erythrocytes are biconcave
discoid-shaped cells.
The major differences are in the size of the red
blood cells and the degree of central pallor.
Listed from largest to smallest in size are :
Dog
Cat
Horse
Cow
Sheep
Goat
Erythrocytes
• Maturation of a RBC.
• Only occurs in the bone marrow of normal
adult animals.
• Occurs in the spleen and liver of the
fetus.
• Maturation time usually takes 5 days.
• Regulated by erythropoietin (EPO) which
is increased in the presence of hypoxia.
• In most species, the kidney is the sensor
organ and major site of EPO.
Red Blood Cells (Erythrocytes)
• No nucleus due to have to fold and
squeeze through tight spaces.
• Normocytes- cells look normal
Erythrocyte Life Span
•
•
•
•
•
•
Dog- 110 days
Cat- 70 days
Cow- 160 days
Horse- 145 days
Man- 120 days
Mouse- 30 days
Erythrocyte Life Span
• Stem Cell → Rubriblast → Prorubricyte →
Metarubricyte → Rubricyte → Reticulocyte
→ RBC
• Rubricyte- nucleated RBC releases in
severe anemia.
• No more mitotic division takes place
after this stage.
• .
Normal Erythrocytes
• Morphologic features vary from species to
species
• Dogs: Biconcave disk shape with
central pallor
• Cats: Generally round with little central
pallor.
• Birds: Contain a nucleated RBC
Categories of Erythrocyte
Characteristics
1. Cell arrangement on the
blood film.
2. Size
3. Color
4. Shape.
5. Presence of structures on
erythrocytes
Cell Arrangement on Blood Film
• Rouleaux
• Rouleaux formation
is a group of
erythrocytes in
stacks. This can be
a sign of increased
fibrinogen or
globulin
concentration
secondary to
inflammation.
• It can also be an
artifact seen in blood
that is held too long
before preparing the
blood slide or in
blood that has been
refrigerated.
Can be a normal
occurence!!
Most Commonly Seen in Horses!!!
Cell Arrangement Continued
• Agglutination
• Agglutination, which
appears as rouleaux,
occurs in immunemediated disorders.
An antibody coats the
cell causing bridging
or clumping.
• If you add a drop of
saline to a drop of
blood rouleaux
formation will disperse
and agglutination will
not
Size
• Terms:
• Anisocytosis:
• Variations in size
• Can indicate anemia
• Macrocytosis:
• Larger than normal cell size
• Liver disease or Vitamin B12
deficiency
• Microcytosis:
• Smaller than normal cell size
• Iron deficiency
MCV
• Mean Corpuscular Volume
• Describes cells as normocytic,
microcytic, or macrocytic. Calculates
the average volume of rbc’s.
• MCV=(Hematocrit x 10)/RBC count in
millions
• Normal: 66-77
Color
• Polychromasia (polychromatophilic):
•
Polychromatic erythrocytes exhibit a bluish tint. The tint is
due to a small amount nucleus retained in the cytoplasm.
These are young cells and may appear as a reticulocyte
• Hypochromasia:
•
•
is a decrease in color, due to a decreased staining intensity
caused by insufficient hemoglobin within the cell.
Iron deficiency is the most common cause.
• Hyperchromasia (hyperchromatophilic):
•
refers to cell that appears darker than normal cells. This
gives the appearance that the cell is over saturated with
hemoglobin. The erythrocyte has a fixed maximum
capacity for hemoglobin and over saturation can NOT
occur.
MCHC
• Mean Corpuscular Hemoglobin
Concentration
• describes cells as normochromatic or
hypochromatic.
• MCHC= (Hgb)/(Hct) x 100
• Normal is 31-36%
• (we will come back to this calculation
again)
Polychromasia
Hypochromasia
Usually associated with
iron deficiency.
Especially in Llamas
Hypochromasia continued
• Hypochromatic should be differentiated
from cells with the center “punched out”. A
punched out appearance can be an artifact
due to improper smear technique
Hyperchroasia
Shape
• Poikilocytosis
• Poikilocytosis is a major deviation in the
normal shape of the erythrocyte. The
term poikilocytosis is an umbrella term
that is used for any and all abnormally
shaped erythrocytes and does not
suggest a specific diagnosis
Schistocytes (Fragmented Cells)
• Also known as poikilocytes.
• RBC’s with abnormal shape.
• Formed as a result of shearing of the cell by fibrin
strands. This occurs when red blood cells rapidly pass
through microvasculature that is lined or meshed with
strands.
• They are observed in fragmentation hemolysis caused by
DIC, vascular neoplasia, endocarditis, and possibly iron
deficiency anemia.
Acanthocytes (Spur Cells)
• The term acanthocyte is derived from
the Greek word “acanthi” meaning
“thorn” Acanthocytes are cells with five
to ten irregular, blunt, finger-like
projections.
• The projections with vary in width,
length and surface distribution. These
cells are seen in animals with altered
lipid metabolism such as cats with
hepatic lipidosis or dogs with liver
disease.
Acanthocytes Continued
Echinoctyes (Burr Cell)
• Echinocytes have multiple, small, delicate
regular shaped spines evenly distributed
around the cell and are indistinguishable
from artificially crenated cells.
Echinoctyes Continued
• Echinocyte formation can
be artificial, often seen with
slow drying blood films or if
the EDTA tube was
underfilled. This artifact is
then termed crenation.
• Echinocytes have been
associated with renal
disease, lymphosarcoma
and rattlesnake bites in
dogs.
• They can be seen after
exercise in horses.
Crenation
• Identified as the presence of many
irregular membrane projections involving
most RBC’s.
• It is usually an artifact due to slow drying
of the blood film.
• Commonly observed in pig blood but can
be seen in any species.
Drepanocytes (Sickle cell)
• These cells are crescent
shaped with pointed ends.
• Drepanocytes are often
seen in normal blood of
deer and goats. It is
thought to be a result of low
oxygen tension.
Drepanocyte cont’d
Keratocyte (Helmet/Blister Cells)
• Also called blister cells or bite cells.
Keratocytes are associated with trauma
especially cellular damage from contact
with fibrin strands.
Prekeratocytes
• Cells with pseudovacuoles are called
blister cells or pre-keratocytes.
Spherocytes
• Cells have a spheroid shape instead of the usual
biconcave disk shape.
• Have reduced cell membrane and are
hypochromatic.
• Seen most frequently in autoimmune hemolytic
anemia (AIHA). When WBC partially remove
antibody-coated membranes.
• Usually seen in dogs.
Stomatocytes
• The appearance of stomatocytes with their
oval or rectangular central pallor has been
compared to a smiling face, a fish mouth,
and a coin slot.
• Stomatocytes are associated with an
hereditary condition but are also seen in
liver disease, acute alcoholism (humans),
and electrolyte imbalances.
Anulocytes
• These are bowl shaped erythrocytes that
form as a loss of membrane flexibility that
does not allow the cell to return to a
normal shape after passing through a
capillary. They can occur due to lowed
hemoglobin concentration or as an artifact.
Dacryocytes (tear drop cells)
• These tear drop shaped cells are seen in
myeloproliferative diseases. These cells
can be produced as an artifact but can be
identified by the direction of their tail
• Dacryocytes produced as an artifact have
their tails pointing in the same direction.
Target Cells and Folded Cells
aka Codocytes
• Two types of cells observed mainly in
dogs.
• Represent cells with an outfolding of the
rbc membrane.
• The cell membrane is thin and flimsy.
• Can be associated with liver dz and
reticulocytosis.
Target Cells (Mexican Hat Cells)
• Also called codocytes
• Thin, bell-shaped cells
• Centrally stained area
• May be seen as
artifacts when smears
made in high humidity
or if blown dry.
Can indicate liver disease
or hemoglobinopathies.
Target Cells
Howell-Jolly Bodies
•
•
•
•
Nuclear remnants observed in young erythrocytes.
Often observed in cats and horses.
Can be seen in regenerative anemic animals.
Also may be seen with splenic disease or in an animal
with the spleen removed.
Nucleated Red Blood Cells
(NRBC’s)
• Nucleated red blood cells (NRBC) usually
represent early release of immature red
blood cells during anemia.
• These are nuclear remnants seen in young
erythrocytes during a response to anemia.
Nucleated Erythrocytes
• Metarubricytes are sometimes found in the
peripheral blood.
• Usually observed with regenerative
anemias.
• May be found in non-regenerative states
such as lead poisoning and hypoxia.
Basophilic Stippling
• Observed in RBC’s that contain abnormal
aggregation of RNA.
• Can be observed in cases of heavy metal
poisoning with non-regenerative anemias
or intense erythrogenesis in dogs, cats,
and ruminants.
Heinz Bodies
• Particles of denature hemoglobin protien.
• They stain with new methylene blue and appear as
colorless bumps with quick stain.
• May be caused by oxidant drugs and chemicals.
• Normal cat blood may have 2-3%.
• Spleen recognizes as abnormal and starts to lyse
the cells.
Reticulocytes
• These are immature red blood cells that
contain organelles (ribosomes) that are
lost in the mature cell. Cats have two
forms of reticulocytes.
• The aggregate form contains large
clumps of reticulum.
• The punctate form, unique to cats,
contains two to eight singular granules
Reticulocytes in the Peripheral
Blood
• Non-nucleated cell containing RNA which
can be easily seen when stained with
methylene blue.
• Hallmark of erythrocyte regenerative
response.