Morphology of RBCs

1. Morphology of blood cells
Hematology Unit
Menoufia University

2.  Blood films should be examined in a systematic manner:
1. Patient identification should be checked and confirmed, and the
microscope slide matched with the corresponding CBC report.
2. The sex and age of the patient should be noted.
3. The film should be examined macroscopically to confirm adequate
spreading and to look for any unusual spreading or staining
characteristics.
4. The film should be examined microscopically.
 Examination should take place first under a low power (×10
objective) → with a high power (× 40 objective) → with oil
immersion (× 100 objective).
Examining the blood film

3. Blood film of a patient with multiple
myeloma (left) compared with another blood
film stained in the same batch (right). The
deeper blue staining occurs due to high
concentration of immunoglobulin leads to
increased uptake of the basic component of
the stain.

4.  On placing a film under the microscope, the first decision to be
made is whether or not it is suitable for further examination.
 It is unwise to give an opinion on an inadequate blood film.

5.  The use of low power examination:
 allows rapid scanning of a large part of the film
 facilitates the detection of abnormal cells when they
are present at a low frequency.
 useful in the appreciation of rouleaux and red cell
agglutination.
 Examination of the blood film must also include
examination of the edges and the tail, since large
abnormal cells and clumps of cells are often
distributed in these areas.
 Platelet aggregates which may cause the platelet
count to be falsely low, if present, are often found
in the tail of the film.
Platelet aggregate in a blood
film

6.  The film appearances should be compared with the CBC report.
 If the CBC and the film are inconsistent with each other, then the
blood specimen should be inspected and the CBC – and if necessary
the film – should be repeated.
 Such discrepancies may be due to:
(i) a poorly mixed or partly clotted specimen.
(ii) a specimen that is too small so that the instrument has aspirated
an inadequate volume; or
(iii) the blood film and CBC being derived from different blood
specimens.
 If such technical errors are eliminated, discrepancy may be due to
an abnormality in the specimen such as the presence of a cold
agglutinin.

7. Erythrocytes

8.  The majority of normal red cells or erythrocytes are disciform in shape.
 On a stained peripheral blood film they are approximately circular in
outline and show only minor variations in shape and moderate variations in
size .
 In the area of a film where cells form a monolayer, a paler central area
occupies approximately the middle third of the cell.
Blood film of a healthy subject showing
normal red cells. The red cells show
little variation in size and shape.

9.  Anisocytosis is an increase in the variability of erythrocyte size beyond
that which is observed in a normal healthy subject.
 Anisocytosis is a common, non‐specific abnormality in hematological
disorders.
 In automated instrument counts, an increase in RDW is indicative of
anisocytosis.
 Poikilocytosis is a state in which there is an increased proportion of cells of
abnormal shape.
 Poikilocytosis is a common, often non‐specific abnormality in many
hematological disorders.
 It may result from the production of abnormal cells by the bone marrow or
from damage to normal cells after release into the blood‐stream.

10. Blood film of a patient with pernicious anemia. Shows
marked anisocytosis, moderate poikilocytosis (including
oval macrocytes and teardrop cells) and a megaloblast.

11.  Microcytosis is a decrease in the size of the erythrocytes.
 The nucleus of a small lymphocyte, is a useful guide to the size of a red
cell.
 Microcytosis may be general or there may be a population of small red
cells. If all or most of the red cells are small there is a reduction in the
MCV, but a small population of microcytes can be present without the
MCV falling below the reference range.
 Macrocytosis is an increase in the size of erythrocytes.
 The erythrocytes of neonates show a considerable degree of macrocytosis.
 A slight degree of macrocytosis is also seen as a physiological feature of
pregnancy.
 Macrocytosis may be a generalized change, and so the MCV will be raised,
or it may affect only a proportion of RBCs.

12.  Hypochromia is a reduction of the staining of the red cell; there is an
increase in central pallor, which occupies more than the normal
approximate one‐third of the red cell diameter.
 Hypochromia may be general or there may be a population of
hypochromic cells.
 Hyperchromia: ×××

13. Microcytic hypochromic anemia Macrocytosis associated with liver
disease

14.  Dimorphism indicates the presence of two distinct populations of red cells.
 The term is most often applied when there is one population of
hypochromic, microcytic cells and another population of normochromic
cells, the latter being either normocytic or macrocytic.
A dimorphic peripheral blood
film

15.  Polychromasia means that some of the red cells stain shades of bluish
grey, these are reticulocytes.
 On average, in patient samples, the reticulocyte count is about double the
visual estimate of polychromatic cells. This is because only the most
immature reticulocytes are polychromatic.

16.  Spherocytosis: Spherocytes are cells that
are more spheroidal (i.e. less disc-like)
than normal red cells but maintain a regular
outline. Their diameter is less and their
thickness is greater than normal.
 In a stained blood film, spherocytes lack
the normal central pallor.
 In examining a blood film for the presence
of spherocytes it is important to examine
that part of the film where the cells are just
touching, since normal cells may lack
central pallor near the tail of the film.

17.  Irregularly contracted cells are cells that lack central pallor and appear
smaller and denser than normal erythrocytes without being as regular in
shape as spherocytes.
 Irregularly contracted cells are formed when there is oxidant damage to
erythrocytes, or damage to red cell membranes by precipitation of unstable
hemoglobin.
Irregularly contracted cells
(arrows), target cells and
hypochromia in a patient
who is a compound
heterozygote for Hb C and
severe beta+ thalassemia.

18.  Elliptocytosis and ovalocytosis : Elliptocytosis indicates the presence of
increased numbers of elliptocytes and ovalocytosis the presence of
increased numbers of ovalocytes.
 elliptocyte is a cell with a long axis more than twice its short axis while
ovalocyte is a cell with the long axis less than twice its short axis.
 When elliptocytes or ovalocytes are numerous and are the dominant
abnormality it is likely that the patient has an inherited abnormality
affecting the red cell cytoskeleton, such as hereditary elliptocytosis.
Elliptocytosis Ovalocytosis

19.  Teardrop cells occur when there is bone marrow fibrosis or severe
dyserythropoiesis and also in some hemolytic anemias.
 They are particularly seen in megaloblastic anemia, thalassemia major and
myelofibrosis.
Blood film of a patient with
primary myelofibrosis showing
teardrop poikilocytes

20.  Target cells have an area of increased staining, which appears in the
middle of the area of central pallor.
 Target cells are formed as a consequence of there being redundant
membrane in relation to the volume of the cytoplasm.
 Target cells may be microcytic, normocytic or macrocytic, depending on
the underlying abnormality and the mechanism of their formation.
 Target cells are much less numerous in iron deficiency than in thalassemia.
Blood film of a hematologically normal
patient who has had a splenectomy,
showing target cells and a Howell–Jolly
body.

21.  Stomatocytes are cells that, on a stained blood film, have a central linear
slit or stoma.
 In hereditary spherocytosis and autoimmune hemolytic anemia, progressive
loss of membrane leads to formation of stomatocytes and spherocytes.
 The commonest acquired cause of stomatocytosis is alcohol excess and
alcoholic liver disease; in these cases there is often associated
macrocytosis.
Blood film in hereditary stomatocytosis
showing stomatocytes.

22.  Sickle cells : A sickle cell is a very specific type of cell that is confined to
sickle cell anemia and other forms of sickle cell disease.
 Sickle cells are crescent‐ or sickle‐shaped with pointed ends.
 The blood film in sickle cell anemia may also show boat‐ or oat‐shaped
cells that are not pathognomonic for the presence of hemoglobin S but are
highly suggestive.
Blood film of a patient with sickle
cell anemia showing sickle cells and
boat-shaped cells.

23. Too much information

24.  Spiculated cells : divided into echinocytes, acanthocytes, keratocytes and
schistocytes.
• Echinocytes (Burr cells): erythrocytes that have lost their disc shape and
are covered with 10–30 short blunt spicules of fairly regular form.
 Some causes of echinocytosis.
• Storage artefact ‘crenation’.
• Liver disease, particularly with co‐existing renal failure.
• Hemolytic–uremic syndrome.
Echinocytes in the peripheral
blood film of a patient with chronic
renal failure

25. • Acanthocytes are cells of approximately spherical shape bearing between 2
and 20 spicules that are of unequal length and distributed irregularly over
the red cell surface. Some of the spicules have club‐shaped rather than
pointed ends.
Numerous acanthocytes in the blood
film of a patient with
abetalipoproteinaemia

27. • Keratocytes (horned cells) have pairs
of spicules, usually either one pair or
two pairs.
 They may result from removal of a
Heinz body (by the pitting action of the
spleen) or from mechanical damage.
 The terms ‘helmet cell’ and ‘bite cell’
have sometimes been used to describe
keratocytes.
 They have been observed in
microangiopathic hemolytic anemia, in
disseminated intravascular coagulation
and in renal disease, e.g.
glomerulonephritis, uremia and
following renal transplantation.
Keratocytes

28. • Schistocytes are fragments of red cells.
 In healthy adult subjects they do not exceed 0.2% of red cells, but in
neonates they may be up to 1.9% and in premature neonates up to
5.5%.
 Schistocytes are formed either by fragmentation of abnormal cells or
following mechanical, toxin‐ or heat‐induced damage of previously
normal cells.
 When resultant on mechanical damage, schistocytes often coexist
with keratocytes.
 Many schistocytes are spiculated. Others have been left with too
little membrane for their cytoplasmic volume and therefore have
formed microspherocytes (spheroschistocytes).
 The commonest causes of schistocyte formation are
microangiopathic and mechanical hemolytic anemias.

29.  ICSH Recommendations for Microscopic Identification of Schistocytes :
1.Schistocytes should be evaluated on PB smears and estimated as a
percentage after counting at least 1000 red blood cells.
2. A schistocyte count should be requested and carried out when a diagnosis
of thrombotic microangiopathies is suspected, usually in patients with
thrombocytopenia.
3. Schistocytes should be identified by specific positive morphological
criteria. Schistocytes are always smaller than intact red cells and can have the
shape of fragments with sharp angles and straight borders, small crescents,
helmet cells, keratocytes, or microspherocytes*.
* Microspherocytes only in the presence of other mentioned RBC shapes.

30. 4. A schistocyte count should be considered clinically meaningful if
schistocytes represent the main morphological red blood cells abnormality in
the smear.
5. A robust morphological indication for the diagnosis of thrombotic
microangiopathic anemia in adults should be recognized when the percentage
of schistocytes is above 1%.
6. Fragmented red cell enumeration by automated blood cell counters should
be considered a useful complement to microscopic evaluation, as it provides
rapid results with a high predictive value of negative samples. A microscope
check is needed for positive and macrocytic samples.†
†Macrocytic samples are at risk of underestimation or absence of flag (‘false-
negative’ test).

31. keratocyte (upper arrow) and helmet
cell (lower arrow), close to a
polychromatophilic erythrocyte in the left
lower corner.
a triangle schistocyte
(arrow) with a
helmet cell on the
upper right.
two
microspherocytes
(arrows)

32. blood film of a patient with
microangiopathic hemolytic. Shows one
very dense microspherocyte and other
red cell fragments.
blood film of a patient with
microangiopathic hemolytic anemia.
Shows numerous bizarrely shaped red
cell fragments.

33. PB smear with comment
on schistocytes

34. Inclusions in erythrocytes
(1) Howell–Jolly bodies : are medium‐sized, round, cytoplasmic red cell
inclusions that have the same staining characteristics as a nucleus and can be
demonstrated to be composed of deoxyribonucleic acid (DNA).
 A Howell–Jolly body is a fragment of nuclear material.
 They appear in the blood following splenectomy .
 They can be a normal finding in neonates (in whom the spleen is
functionally immature).
 The rate of formation of Howell–Jolly bodies is increased in megaloblastic
anemias and, if the patient is also hyposplenic, large numbers of Howell–
Jolly bodies will be seen in the peripheral blood.

35. (2) Basophilic stippling or punctate basophilia describes the presence in
erythrocytes of considerable numbers of small basophilic inclusions that are
dispersed through the erythrocyte cytoplasm and can be demonstrated to be RNA.
 Punctate basophilia seen in:
 thalassemia minor, thalassemia major
 megaloblastic anemia
 unstable hemoglobins
 hemolytic anemia
 dyserythropoietic states in general (including congenital dyserythropoietic
anemia, sideroblastic anemia, and primary myelofibrosis)
 liver disease and poisoning by heavy metals such as lead.
 Hereditary deficiency of pyrimidine 5’‐nucleotidase ,an enzyme that is required
for RNA degradation. Inhibition of this enzyme may also be responsible for the
prominent basophilic stippling in some patients with lead poisoning.

36. Basophilic stippling

37. (3) Pappenheimer bodies are basophilic inclusions that may be present in
small numbers in erythrocytes; they often occur in small clusters towards
the periphery of the cell and can be demonstrated to contain iron.
 They stain on a Romanowsky stain.
 A cell containing Pappenheimer bodies is a siderocyte.
 Following splenectomy in a hematologically normal subject, small
numbers of Pappenheimer bodies appear, these being ferritin
aggregates.
 In pathological conditions, such as lead poisoning or sideroblastic
anemia, Pappenheimer bodies can also represent iron‐laden
mitochondria. If the patient has also had a splenectomy they will be
present in much larger numbers.

39. (4) Heinz bodies are clumps of irreversibly denatured hemoglobin
attached to the erythrocyte cell membrane.
 Heinz bodies are indicative of oxidative injury to the erythrocyte.
 They are seen in hemolytic anemia due to unstable hemoglobins,
exposure to oxidizing drugs, chemical poisoning, G6PD
deficiency.
 Supravital staining is required to see Heinz bodies.

40. Bite cell

41.  Circulating nucleated red blood cells
 Except in the neonatal period and occasionally in pregnancy, the presence
of NRBC in the peripheral blood is abnormal, generally indicating
hyperplastic erythropoiesis or bone marrow infiltration.
 If both NRBC and granulocyte precursors are present the film is described
as leucoerythroblastic.
 NRBC in the peripheral blood may be morphologically abnormal; e.g. they
may be megaloblastic or show the features of iron deficient.

42.  Red cell agglutination, rouleaux formation
 Red cell agglutinates are irregular clumps of cells.
 Rouleaux are stacks of erythrocytes resembling a pile of coins.
Red cell agglutination Rouleaux formation