6. Transfer the buffy coat to Wintrobe’s hematocrit tube and centrifuge again for 5–10 minutes. 7. Transfer the buffy coat and an equal volume of plasma to a small tube, mix well, and prepare smears. Dry rapidly and stain with Giemsa’s or Leishman’s
¾ Recovery from agranulocytosis.
¾ Iron deficiency anemia (some cases)
¾ Hemolytic and toxic anemias of long standing
Morphologic forms of Lymphocytes
(Also called stress lymphocytes, Downey type cells, or
1. These are small atypical cells that appear in viral
diseases such as mononucleosis, viral hepatitis, viral
pneumonia, and viral upper respiratory tract infections.
2. These may also be found in numerous nonviral
a. Fungoid and protozoid nonviral conditions
d. After transfusions and tissue graft.
3. When seen in stress response, these are called stress
4. May be found in apparently healthy children.
5. Up to 10% of all lymphocytes, can be considered
a. Lymphocyte cells that may be seen in macroglobulinemia.
b. Turk cells and Reider cells that are seen in acute
c. Vacuolated lymphocytes that are seen in lipidosis.
2. Culturing of lymphocytes in laboratory:
a. Stimulates small lymphocytes to transform into
large atypical cells which produce immunoglobulin.
b. Transformation response is impaired in culturing
of lymphocytes from patients with:
c. Transformation response increased in sarcoidosis.
3. Other uses of transformation test are to determine
histocompatibility of recipient and donor for tissue
a. Lymphocytes from donor not related to recipient
stimulate the production of up to 3% of transformed lymphocytes in recipient.
b. Lymphocytes from sibling react less strongly.
c. No reaction occurs on cultures from fraternal
262 Concise Book of Medical Laboratory Technology: Methods and Interpretations Arneth Count
Neutrophils can be divided into five main groups according
to the number of lobes in their nuclei.
Group 1 One lobe, even if it shows indentation and
thinning out at one or more places.
Group 2 Two lobes connected by one thin filament.
Group 3 Three lobes connected by two thin filaments.
Group 4 Four lobes connected by three thin filaments.
Group 5 Five or more lobes connected by four or more
Using the above-mentioned classification, count 100
neutrophils and the number in each group is to be
expressed as percentage. Usual normal values:
Interpretation: In acute infections, there is rapid turnover
of neutrophils and in the process younger neutrophils with
lesser number of lobes are released into circulation, thus
increasing the number of cells in groups 1 and 2 (shift to
In macrocytic megaloblastic anemias, the neutrophil
production rate is slow, hence cells with hypersegmented
nuclei are released into circulation (shift to the right).
However, Arneth count is now no longer in use.
The percentage of cells in groups 1, 2 and ½ of 3 is about
60 (normal range 51–65). Only 2–5% fall in group 1.
Neutropenia and Agranulocytosis
¾ Adrenocorticosteroid therapy (transient)
¾ neutrophil leukocytosis or leukemoid reaction associated
• Following prolonged corticosteroid therapy.
Excessive leukocytic response to a stimulus and/or
immature cell spilling over in peripheral blood.
¾ Vascular thrombosis and infarction
¾ Marrow replacement and myeloid metaplasia.
¾ Systemic lupus erythematosus.
Neutropenia and Agranulocytosis
Neutropenia is the reduction in number of circulating
neutrophils below 2500 cells/cu mm.
Blood Picture of Drug-induced Neutropenia
¾ Neutropenia with no anemia or thrombocytopenia
¾ In some cases, there may be lymphopenia and
¾ Neutrophils may show toxic and degenerative changes
¾ Absence of granulocytic precursors with normal
erythropoiesis and a normal number of megakaryocytes
¾ Toxic granulation in developing granulocytes
¾ Granulocytic hyperplasia implies recovery.
1. Drugs that cause aplastic anemia.
2. Drugs that induce selective neutropenia:
Antipyretic analgesics—Amidopyrine.
Antithyroid drugs—Thiouracil, methimazole, carbimazole.
Antihistamines—Promethazine, chlorpheniramine,
Tranquillizers and antidepressants —Chlorpromazine,
meprobamate, imipramine, amitri-ptiline, etc.
Antibacterials—Tetracycline, streptomycin, ristocetin,
salazopyrin, sodium methicillin, etc.
Anticoagulants—Phenindione, dicoumarol.
Antituberculars—Isoniazid, PAS, thiacetazone.
Antimalarials—Primaquine, amodiaquin.
Miscellaneous—Procainamide, penicillamine, metronidazole, etc.
1. Chronic idiopathic neutropenia (agranulocytosis)
neutrophil count = 500–2000 cells/cu mm— absolute
3. Marrow aplasia: All causes of aplastic anemia.
4. Due to known cause or myelophthisis
• Folic acid or Vitamin B12 deficiency causes
megaloblastic or macrocytic anemia also.
• Pancreatic exocrine deficiency
• Cyclic neutropenia (familial/sporadic).
Symptomatic neutropenia occurs usually in aplastic
anemia, drug-induced neutropenia, hypersplenism and
idiopathic neutropenia, and acute leukemia.
Laboratory Diagnosis Of Infectious Mononucleosis
¾ There is both absolute and relative lymphocytosis, large
¾ Hemoglobin value and platelet counts are normal
¾ Initial leukopenia due to reduction in number of
neutrophils, the neutrophil alkaline phosphatase count
¾ Lymphocytosis is maximum at about the tenth day
¾ Lymphocytosis (atypical) as described by Downey and
• Type I—Monocytoid lymphocytes
• Type II—Plasmacytoid lymphocytes
• Type III—Blastoid lymphocytes
¾ Wasserman reaction may be positive in 3–10% cases
¾ Latex and particle agglutination tests are also available.
Paul Bunnel Test for Heterophile Antibody
This is based upon the presence of antisheep red cell
hemagglutinins in unusually high titers in the sera of these
patients. It is positive in about 80–90% of cases. It remains
positive for a variable period of time. In addition to the said
antibodies, at least 2 other types of agglutinin for sheep red
cells occur in human serum. They are:
1. An antibody present in low titers in normal persons
and in malignant lymphomas. This antibody is
absorbed by guinea pig kidney but not by ox cells.
2. An antibody occurring following the injection of horse
serum and in serum sickness. This is absorbed both
by guinea pig kidney and ox cells. This antibody of
infectious mononucleosis is not absorbed by guinea
pig kidney but is absorbed by ox cells.
Lupus Erythematosus (LE) Cell/Phenomenon
1. Draw 5–10 mL of venous blood. Place in a 50 mL flask
containing 20–30 glass beads 3–5 mm in diameter or
clear metal paper clips. Swirl or shake gently for 10–15
minutes to defibrinate the blood.
2. Let stand 15 minutes (preferably at 37°C).
3. Transfer blood and a few beads to a test tube or
container and mix on a rotator or by inverting for 30
4. Let stand at room temperature (preferably at 37°C) for
5. Centrifuge at 2000–3000 rpm for 5–10 minutes.
6. Transfer the buffy coat to Wintrobe’s hematocrit tube
and centrifuge again for 5–10 minutes.
7. Transfer the buffy coat and an equal volume of
plasma to a small tube, mix well, and prepare smears.
Dry rapidly and stain with Giemsa’s or Leishman’s
8. Examine smears for clumps of platelets and neutrophilic
leukocytes where the typical LE cell is most likely to
be found. Look for neutrophils containing ingested
homogeneous blue to magenta colored bodies (LE
cell) or a group of neutrophils encircling (garlanding)
It should be differentiated from ‘tart cell’ in which though
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