This document provides information on examining blood smears and grading abnormal red blood cells. It discusses red blood cell terminology and variations in size, shape, staining and inclusions. It describes different abnormal red blood cell morphologies including anisocytosis, poikilocytosis, hypochromia, polychromasia, and inclusions. Grading of abnormalities from few to 4+ is explained. Specific red blood cell changes seen in various hematological conditions are mentioned. Guidelines for examining blood smears under different microscope objectives and estimating white blood cell counts and platelets are provided. Key references on the topic are listed at the end.

1. Assoc.Prof. Pornsri Tantiniti
Faculty of Medical Technology
Rangsit University
Blood smear examination
and grading of RBC

2. ●Explain Rbc terminology and pathogenesis
●Detail red blood cells variations in regards to
size, shape, and color etc.
●Grading abnormal red blood cell in patients
●How to examine blood smear
To know about . . . .

3. Pathogenesis
● Nutritional anemia: Iron deficiency,
Megaloblastic anemia
● Membrane cytoskeletal defect :
Ovalocyte, spherocyte
● Defect metabolite state of rbc :
● E-M pathway (Enz. Deficiency)
● ATP production ( ATP , Na influx),
● Glycolytic Enz.Defect (G6PD Deficiency)

4. Pathogenesis
● Defect in Hb synthesis & ab.Hb
● Thalassemia
● HbE
● Defect in Lipid bilayer membrane
● Acanthocyte in abeta-liproteinemia
● Anemia chronic disease
● Leukemia
● SLE

5. Red Blood Cell Morphology

6. • The blood film
should be examined
in the area where the
red cells are
touching but not
often overlapping.
• At least 10 oil field
• 1 oil field ~ 200 RBC
Standard area

7. Red cell terminology
• Anisocytosis : variation of red cell sizes
• Poikilocytosis : variation of red cell shapes
• Staining
- Normochromic : red cell with normal color
- Normocytic : red cell with normal size
• Dimorphic red cell : presence of two
different populations of red cell (different in
size , Hb.content , shape ) e.g.
Hypo micro rbc VS. Macrocytic red cell

8. Red cell terminology
• Inclusion in rbc
• Distribution
• NRC : nucleated red cell

9. 1. Size
2. Shape
3. Staining
4. Inclusion
5. Distribution
6. NRC
Abnormal Morphology

10. 1. Size
Normal RBC (7 µm) >>>
• Variation in size >>>> Anisocytosis
• Macrocyte and Microcyte in blood
smear

11. Normocyte
• Normal red cells or
erythrocytes show only slight
variation in size and shape.
•Biconcave
•7 µm(6.2-8.2)
•Central pallor 1/3
1. Size

12. 1 Anisocytosis (Variation in size)
Microcyte
•Size <6.2 µm
•MCV < 80 fL
• Microcytosis
Causes
• Thalassemia, sideroblastic
anemia Pyruvate-kinase
deficiency

13. Macrocyte
•Size > 8.2 µm
• MCV > 100 fL
• Macrocytosis
Pathogenesis
- Abnormal erythropoiesis
- Pernicious anemia
- Vit.B12,Folic acid deficiency
- After acute blood loss
- Hemolytic disease of newborn
1 Anisocytosis (Variation in size)

14. Normal RBC >>> Biconcave shape
round shape
• Variation in shape >>>> Poikilocytosis

15. 2.1Poikilocytosis (Variation in shapes)
Ovalocyte or Elliptocyte
• Cigar or egg shaped RBC
• Hb conc. > normal
- Normal < 1%
Causes
-Hereditary elliptocytosis or
Ovalocytosis
- Megaloblastic anemia
- Iron deficiency
- Myelofibrosis

16. ●The macrocyte is oval
instead of round (MCV >
125 fL )
●In most instances is due
to megaloblastic
erythropoiesis (Vitamin
B12 or folate deficiency)
but may be seen with
dyserythropoiesis

17. -Small, spherical cells lacking central pallor
Pathogenesis
-Result from partial removal of RBC
membrane by monocyte-macrophage
system
- Decreased membrane surface
area:cytoplasmic volume ratio
Causes
- Hereditary spherocytosis,
immuno-hemolytic anemia
2. Poikilocytosis or Variation in shape
Spherocyte

18. 2.2 Poikilocytosis
Target cell
-Bull’s eye appearance,Hb
accumulation in center and periphery
with clear interventing
- May be microcyte, nomocyte,
macrocyte.
- Pathogenesis
- Area membrane phospholipids
- Membrane cholesterol
Causes
Liver disorders, iron deficiency,
thalassemia, hemoglobinopathies,

19. 2.3 Poikilocytosis
Teardrop Cell
-Tear drop-shaped cells. Pathogenesis:
pitting function of spleen
Causes
- Thalassemia
- Drug induced “Heinz bodies”
- Extramedullary erythropoiesis
- Myeloid metaplasia
“ Myelofibrosis ”

20. Stomatocyte
-Cup shape,Uniconcave
Causes
- Cationic drugs
Low “ pH “ response
- Liver disease
- Alcohol intake
- Hereditary stomatocytosis
2.4 Poikilocytosis

21. 2.5 Poikilocytosis
Sickle cell
-Elongated and narrow with one or
both ends curved and pointed,
crescent-shaped.
Causes
- Abnormal Hemoglobin S

22. 2.6 Poikilocytosis
Ecchinocytes (Crenated)
Disc shape and are covered with
10-30 short blunt spicules of
regular form
Causes
Storage artifact (EDTA blood,
delayed in making bl. film)

23. Acanthocyte (spur cell)
-Spherical shape with 2-20 spicules and
unequal length.
Finger-like projections
Pathogenesis
Expansion of the outer leaflet of lipid
bilayer
Cholesterol /Phospholipid ratio
Causes
- Alcoholic liver cirrhosis, - Hepatitis
- Hereditary abetalipoproteinemia
2.7 Poikilocytosis

24. - Cell with pairs of spiny 2 or 4-6 regular
projections on surface
- Horned cell
Pathogenesis
Rupture or removal of circular area of
apposed, sealed cellular membrane
projections formed
Causes
- Uremia
- MAHA
- DIC
(Disseminated intravascular coagulation)
Keratocyte (burr cell)
2.8 Poikilocytosis

25. Schistocyte
Triangular or helmet shaped fragments of RBC
with sharp, pointed projections, and irregular
borders
Pathogenesis
Fragmentation from impact with fibrin strands,
walls of diseased vessels, artificial surfaces,
and turbulent blood flow- Physical/Mechanical
stress
Causes
- Drugs
- Chemical
- Thalassemia
- DIC
(Disseminated intravascular coagulation)
2.9 Poikilocytosis

26. Normochromia
Hypochromia
3. Staining
Polyochrom
asia
- Normochromia
- Hypochromia
- Polychromasia

27. Hypochromia
●Central pallor > 1/3,hemoglobin
concentration is decreased
• Hemoglobin concentration decrease
can be caused by decreased amounts
of iron available for hemoglobin
production
• Iron deficiency, thalassemia,
hemoglobinopathies

28. ● Reticulocyte
● Larger than Normal RBC
and purple color
● Normal found 0.2-2%
● Iron deficiency
● Thalassemia
● Hemoglobinopathies
Polyochromasia

29. 4. Inclusion
1. Basophilic Stippling
RNA left over
- Aggregation of rRNA in the
cytoplasm of RBC with Purple-blue
granules
- Lead poisoning
- Pyrimidine-5-nucleotide def.
- Disorders of Hb synthesis
- Hb Constant Spring

30. 4.2 Pappenheimer Bodies
Pappenheimer Bodies
- Granules of ferritin
aggregate in rbc
- Iron+mitochondria+
ribosomes
- Spleenectomy
- Lead poisoning ,
sideroblastic anemia

31. 4.3 Howell-Jolly bodies
DNA left over
• Basophilic nuclear
remnants (clusters of DNA)
in circulating erythrocytes.
• Megaloblastic anemia
• Thalassemia
• Post -splenectomy

32. “Spindle fibers”
microtubules of
mitotic spindle
- Severe anemia
- Dyserythropoiesis
- Splenectomy
4.4 Carbot’s ring

33. 4.5 Malaria

34. 4.6 Heinz bodies & Inclusion bodies
”Supravital stain” unstable
hemoglobin: Hb C
-G-6-PD deficiency
-Inclusion bodies
- Supravital stain
-Unstable hemoglobin: Hb H
(β4) inclusion bodies
Inclusion bodies

35. 5. Distribution
1. Rouleaux Formation
•Hyperproteinemia
•Macroglobulinemia
- Multiple myeloma

36. 5.2 Agglutination
Agglutination
- Antigen - Antibody
- Cold hemagglutination
(AIHA)
- Paroxysmal cold
hemoglobinuria
Agglutination 3+

37. 6. Nucleated rbc
-Immature rbc mostly are
polychromatic normoblast and
orthochromatic normoblast
- report : no of NRC /100 wbc

38. ideal area
Screen with objective 10x for good area

39. - Anisocytosis (abnormal in size)
- Poikilocytosis (abnormal in shape)
- Color
- Red cell inclusions
-Distribution
-NRC

40. Grading:- few ,1+, 2+, 3+ & 4+
Report
few = <10 %
= 11-25 %
= 26-50 %
= 51-75 %
1+
2+
3+
4+
= >75 %
• macroovalocyte,acanthocyte
• teardrop cell, spherocyte
(microspherocyte)
***** If found you must reported

41. 1. Anisocytosis (Abnormal in size)
- Microcyte
- Macrocyte
Sum of microcyte and macrocyte <,= anisocytosis
Example
Anisocytosis 2+ (microcyte 1+, macrocyte some)

42. 4.2 Poikilocytosis (abnormal in shape)
- target, tear drop, schistocyte,ovalocyte etc.
Sum of target, tear drop,schistocyte(at least 3 different
shapes)= poikilocytosis
Example
Poikilocytosis 3+ (target cell 2+,
few spherocytes few burr cells)
Howell Jolly body: found
Few NRC (2 cells/100WBC)

43. 3. Color & Distribution
1. Hypochromia
Normal>>>central pallor = 1/3
1+
2+
3+
4+
------ >
------ >
------ >
------ >
central pallor >1/3 – 1/2
central pallor > 1/2
central pallor >2/3
No No
4.3.2 Polychromasia *****Must report when found 10 oil
fields
0-1 cells/OF
2-3 cells/OF
4-6 cells/OF
few ----- > polychromasia
1+ ------ > polychromasia
2+ ------ > polychromasia
3+ ------ > polychromasia
4+
------ > polychromasia
7-12 cells/OF
> 12 cells/OF

44. Distribution => FOUND, SEEN
Rouleaux formation and Autoagglutination
1+ = 11-25 %
2+ = 26-50 %
3+ = 51-75 %
4+ = >75 %
4.4 Red cell inclusions
*****Must report when found****
“ Present or found “

45. Few target cell , few burr cell

46. Ovalocytosis 3+

47. Normal
Microcyte
Macrocyte
Anisocytosis 2+ (macrocyte 2+ , few microcyte)
Hypochromia 2+

48. Anemia, spherocyte 3+, polychromasia 2+

50. Schistocyte 1+, polychromasia 2+

51. Basophilic stippling found

52. Howell Jolly bodies found

53. Rouleaux formation 1+

54. Malaria P.vivax , schiszont found

55. 10 x Objective
● Blood smear
● 10x estimation (Low power objective lens)
● Cell distribution, ideal area (std area)
● RBC agglutination, Rouleaux formation
● Scan abnormal cells (granulocyte precursors,
NRC)
Blood smear

56. 40 x Objective
● Variation of Rbc size,shape, staining
● Scan toxic granules, Howell jolly bodies
● Pappenheimer body
Blood smear

57. 40 x Estimate
WBC / HP ~ WBC x 109 / L
2 – 4 4.0 – 7.0
4 – 6 7.0 – 10.0
6 – 10 10.0 – 13.0
10 – 20 13.0 – 18.0
Blood smear

58. 100x Objective oil immersion
● Differential WBC
● Estimate platelet
● Rbc morphology report
Blood smear

59. Platelet
●Estimate platelet : AVERAGE 10 consecutive
oil fields
●Adequate 5-25 /OF
●Slightly decreased 3-4 /OF
●Moderately decreased 1-2 /OF
●Markedly decreased 0-1 /OF
- Pale stain & giant platelet. Must be reported

60. 1. BAIN J B, BLOOD CELLS A PRACTICAL GUIDE.
BLACKWELL SCIENCE, 2002
2. นันทรัตน โฆมานะสิน, นพมาศ เข็มทองหลาง, มณเฑียร
พันธุ เมธากุล. การทดสอบพื้นฐานทางห องปฏิบัติการโลหิต
วิทยา ภาค วิชาจุลทรรศน คลินิก คณะเทคนิคการแพทย
มหาวิทยาลัย ขอนแก น. พิมพคร ััังที่ 2. หจก. โรงพิมพคล
ัังนานาวิทยา, 2544.
3. สุภินันท สเป ค-สายเชื้อ. ภาพสีประกอบ โลหิตวิทยา.พิมพ
ครั้งที่ 4.
กรุงเทพ: เอช ที พี เพรส, 2544.