Hema 2 WBC
Hema 2 WBC
Hema 2 WBC
LEUKOCYTES
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Content of Neutrophil Granules
Primary Granules / Secondary Granules / Tertiary Granules Secretory granules
Azurophilic / Specific Granules
Non-specific Granules
Myeloperoxidase Lactoferrin Gelatinase (ONLY) CD11b/CD18
Acid hydrolase Lysozyme CD10, CD13, CD14, CD16
Acid phosphatase Collagenase Cytochrome b558
β-glucuronidase Aminopeptidase Complement 1q receptor
Arylsulfatase Plasminogen activator
Elastase
Neutrophils
• Once in the peripheral blood, neutrophils are divided into:
o Circulating neutrophil pool (CNP) 50%
o Marginated neutrophil pool (MNP) 50%
• H-L in the blood: 7 hrs. / Lifespan: 2-3 days
• Neutrophilia is an absolute increase in neutrophils > 7 x 109/L in adults or 8.5 x 109/L in children.
• Neutropenia is a decrease in the ANC < 2 x 109/L in adults.
• The term leukemoid reaction refers to a neutrophilic leukocytosis > 50 x 109/L with a shift to the left (ex:
Metamyelocyte; Band/Stab).
• Leukemoid Reaction is usually caused by acute / chronic infections, metabolic disease, inflammation or as part
of an inflammatory response to malignancy.
REFERENCE RANGE:
WBC
Newborns: 10-30 x 109/L
Adults: 3.6-10.6 x 109/L
DIFFERENTIAL
RELATIVE COUNT
Neutrophil: 50-70%
Lymphocytes: 18-42%
Monocytes: 2-11%
Eosinophils: 1-3%
Basophils: 0-2%
ABSOLUTE COUNT
Neutrophils: 1.7-7.5 x 109/L
Lymphocytes: 1.0-3.2 x 109/L
Monocytes: 0.1-1.3 x 109/L
Eosinophils: 0-0.3 x 109/L
Basophils: 0-0.2 x 109/L
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Causes of Neutrophilia
• Emotional stress • Vasculitis • Leukocyte adhesion deficiency
• Strenuous exercise • Colitis • Familial cold urticaria
• Trauma/injury • Autoimmune disease • Hereditary neutrophilia
• Pregnancy: labor and delivery • Acute hemorrhage • Leukemia
• Eclampsia • Chronic blood loss • Chronic myeloproliferative neoplasm
• Surgery • Hemolysis • Steroids
• Infections: bacterial, some viral • Smoking • Lithium
• Burns • Chronic blood loss • Colony-stimulating factors (granulo-
cyte CSF)
• Myocardial infarction • Metabolic ketoacidosis
• Cortisol
• Pancreatitis • Uremia
• Epinephrine
Drugs Other
• Anticancer • Levamisole (adulterated cocaine)
• Analgesics/antiinflammatories, acetaminophen, aspirin, • Immunologic
diclofenac, ibupro- fen, indomethacin, linezolid, naproxen,
sulfasalazine Radiation
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Eosinophils
• Eosinophil development is similar to neutrophils.
• First maturation phase that can be identified is myelocyte stage (reddish-orange 2° granules)
• Mature eosinophils have bi-lobed nucleus
• H-L in the blood: 18 hrs. / Lifespan: 2-5 days
• Counteracts the factors released by mast cells
• Eosinophilia is associated with helminthic infections and allergic reactions.
• Specific Granules contain: 1. Major Basic Protein; “Anti-helminthic properties”
2. Eosinophil Cationic Protein
• Charcot-Leyden Crystals (STOOL/SPUTUM)
Hexagonal bipyramidal crystals composed of Lysophospholipase
Present in the cytoplasm of eosinophils in prolonged eosinophilic inflammation
EOSINOPHILIA EOSINOPΕΝΙΑ
Absolute eosinophil count > 0.4 x 109/L Absolute eosinophil count 0.09 X 10/L
Causes: Causes:
Helminthic infections BM hypoplasia
allergic reactions (asthma, autoimmune disorders
rhinitis, urticaria) steroid therapy
scarlet fever stress
primary biliary cirrhosis sepsis
Hepatitis acute inflammatory states
HIV
autoimmune disorders
drug reactions
Basophils
• Partially lobulated nucleus, obscured by its blue-black granules
• Closely related to Mast cells (effector cell in type 1 hypersensitivity) (involved in immediate hypersensitivity
reactions.
• Basophilia is associated with CML, allergic rhinitis, chronic infections, hypothyroidism
• Specific granules contain: 1. Histamine
2. Heparan Sulfate
3. Leukotrienes
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MONONUCLEAR CELLS
Lymphocytes
• Subsets:
o T cells – 60-70 %
o B cells – 10-20 %
o NK cells – 10-15 %
• High N:C ratio with scanty cytoplasm
• They are not end-stage cells; they are still capable of mitosis when they are stimulated
• Lymphocytosis is usually accompanied by changes in lymphocyte morphology and are associated mainly with
viral infections such as IM, CMV infection, Hepatitis, HIV, Chickenpox, Herpes, Influenza.
• Lymphocytosis
Children: absolute lymphocyte count > 10 x 109/L
Adults: absolute lymphocyte count > 5 x 109/L
• Lymphocytopenia
Children: absolute lymphocyte count < 2 x 109/L
Adults: absolute lymphocyte count < 1 x 109/L
Surface CD2 - “E-rosette” receptor (Has the CD19 - part of B cell coreceptor. CD16 - receptor for IgG
Markers highest affinity to steep RBC) CD20 - found on all stages of B cells. CD56 - no known
CD3 - part of T-cell antigen receptor CD21 - receptor for C3d and EBV. function
CD4 - Helper T-cell / Treg cell CD22 - found mostly on mature B
CD5/7 - early stages of T cells cells.
CD8 - Cytotoxic T-cell CD10 - early stages of B cells/CALLA.
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Monocytes
• Maturation stages: Monoblast, promonocyte, monocyte
• Appearance on stained smear:
o Nucleus is may be round, oval, kidney shaped or horse-shoe
shaped
o Has stringy or lace-like chromatin pattern
o Cytoplasm is bluish gray with fine azure granules (“ground-
glass” appearance)
o Cytoplasmic and nuclear vacuoles may also be present
MONOCYTOSIS MONOCYTΟΡΕΝΙΑ
Tuberculosis Aplastic anemia
Brucellosis Chemotherapy
Leishmaniasis Steroid therapy
Subacute bacterial endocarditis Hemodialysis
Malaria EBV infection
SLE, RA Hairy cell leukemia
Inflammatory bowel disease
Myositis, Sarcoidosis
Malignancy
Hodgkin disease
Myelodysplastic/myeloproliferative neoplasms
Acute myeloid leukemia
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Non-Malignant Leukocyte Disorders
2. Wiskott-Aldrich Syndrome
• rare X-linked disease caused by one of more than 400 mutations in the WAS gene
• T cells are decreased; B cells, T cells and NK cells, neutrophils and monocytes are dysfunctional
• There is a risk of bleeding due to thrombocytopenia and small, abnormal PLATELETS.
5. Chédiak-Higashi Syndrome
• rare autosomal recessive disease of immune dysregulation
• Leukocytes exhibit abnormally large lysosomes, containing fused dysfunctional granules
• Giant lysosomal granules are present in granulocytes, monocytes, and lymphocytes
• Patients often have bleeding issues as a result of abnormal dense granules in platelets
6. Leukocyte Adhesion Disorders
• Rare autosomal recessive inherited conditions
• Inability of neutrophils & monocytes to move from circulation to the site of inflammation (extravasation or
DIAPEDESIS).
• Lab Evaluation:
1. DHR Assay — Dihydrorhodamine; Based on Flow Cytometry.
2. NBT Dye Test — Nitroblue Tetrazolium
8. WHIM Syndrome
• Warts, Hypogammaglobulinemia, Infections, and Myelokathexis syndrome (which causes Neutropenia).
• Defective movement of WBCs between the bone marrow and peripheral blood.
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Morphologic Abnormalities of Leukocytes
2. Hypersegmentation of Neutrophils
• More than 5 nuclear lobes present
• Associated with: Megaloblastic anemia, Myelodysplastic syndrome
3. Alder-Reilly Anomaly
• Autosomal recessive
• Granulocytes have dark staining granules (Reilly bodies) NOT related to infection
• Root cause: Incomplete degradation of mucopolysaccharides
• Neutrophils are functioning normally; can also be present in monocytes and lymphocytes,
• Commonly associated with Hurler syndrome, Hunter syndrome, Sanfilippo syndrome
4. Auer Rods
• Rod-like bodies representing aggregated primary granules
• Reddish-purple granules; Peroxidase (+)
• Observed in blast cells present in AML,
Especially, APL: Acute Promyelocytic Leukemia
• Faggot cells = bundles of Auer rods
5. May-Hegglin Anomaly
• Rare, autosomal dominant disorder
• Triad:
o Dohle-like inclusion bodies
= pale blue inclusion
o Thrombocytopenia
o Giant Platelets MACROTHROMBOCYTOPENIA
• There is disordered production of myosin heavy chain type IIA, which affects megakaryocyte maturation and
platelet fragmentation when shedding from megakaryocytes.
• Patients are usually asymptomatic
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• Observed in severe infections, other toxic conditions, and reactive conditions
2. Dohle Bodies (Neutrophils)
• Round or elongated, pale-blue staining bodies located close to cell membrane
• Cytoplasmic remnants of RNA or free ribosomes
• Found in bacterial infections, burns, pregnancy
3. Flame cell
• variant of plasma cell with irregular cytoplasmic projections (bright purple-red in Wright’s stain)
• These cells contain more immunoglobulin than normal plasma cells
• Associated with Plasma Cell Myeloma / Multiple Myeloma (Refers to the malignancy of plasma cells).
5. Sezary cell
• Lymphocyte with convoluted, cerebriform nucleus (“brain-like”)
• Associated with: 1. Sezary Syndrome
2. Mycoses Fungoides
2. Niemann-Pick Disease
• characterized by an accumulation of fat in cellular lysosomes of vital organs, which impairs function
• deficiency in the enzyme Sphingomyelinase.
• Foam cells and sea-blue histiocytes can be seen in the bone marrow.
o Foam cells have cytoplasm packed with lipid-filled lysosomes (small vacuoles)
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o Sea blue histiocytes are macrophages with lipofuscin-, glycophospholipid-, and sphingomyelin
contained in -cytoplasmic granules, that appear blue with Wright stain.
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LEUKEMIA
• refers to the rapid, clonal proliferation in the bone marrow of lymphoid or myeloid progenitor cells known as
lymphoblasts and myeloblasts.
• development of leukemia is currently believed to be a progression of mutations that give leukemic stem cells a
proliferative advantage and also hinder differentiation
ACUTE CHRONIC
Predominant cell type Precursors / blasts Mature
Onset Sudden Insidious
Symptoms Fever, fatigue, bleeding Variable; non-specific
WBC count Variable Increased
Progression w/o treatment Rapid; weeks to months Slow; months to years
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Major types of AML (Based on WHO System) / Cytogenetics Abnormalities
AML with t (8;21) corresponds to acute myeloid leukemia with neutrophilic maturation;
also known as core binding factor leukemia
AML with inv (16) correspond to acute myelomonocytic leukemia with abnormal eosinophils; eosinophils are
increased and have large basophilic granules
APL with t (15;17) promyeloctes predominate in the BM; Azurophilic granules are abundant;
Multiple Auer Rods are usually present; DIC is common
AML with t (6;9) associated with multilineage dysplasia and basophilia of marrow and blood; shows
variable morphology and granulocytic and/or monocytic differentiation
AML with inv (3) shows multilineage dysplasia, increased atypical megakaryocytes with monolobed or bilobed
nuclei, and normal to increased platelet counts.
AML with t (1;22) presenting as acute megakaryoblastic leukemia with hepatosplenomegaly; acute
leukemia of infants and young children, often reported in girls
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FAB Classification of ALL
L1 L2 L3
Cell size Small Large Large
Chromatin Homogenous Heterogenous Homogenous
Nuclear shape Regular Irregular Regular
Nucleoli Rare Present Prominent
Cytoplasm Scanty (↑ N:C ratio) Moderate Moderate & deeply basophilic
Occurrence CHILDREN ADULTS Leukemic phase of Burkitt’s lymphoma
Cytochemical Stains
AML ALL
MPO (Myeloperoxidase) + -
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Myeloproliferative Neoplasms
Caused by genetic mutations in the stem cell that causes excessive production and accumulation of RBCs,
granulocytes and platelets in the BM, blood and tissues.
• CBC findings:
o Increased Total WBC count (> 50 x 109/L)
▪ Increased in mature forms of granulocytes –
Neutrophils, Eosinophils, Basophils
▪ Myeloblasts are only 1-5%
o Normal or increased platelet count
o Normal or decreased RBCs
Acute myelogenous leukemia – blast / immature cells; >20% blasts, >30% blasts
Both leukemoid reactions & CML has high WBC count; most are mature WBCs
• Must be differentiated from Leukemoid reactions (associated with severe infections) thru NAP cytochemical
staining
o ↓ NAP score = CML
o ↑ or normal NAP score = leukemoid reactions
NAP – neutrophil alkaline phosphatase
LAP – leukocyte alkaline phosphatase
• The neutrophil alkaline phosphatase (NAP) is greatly reduced or absent in > 90% of patients with CML.
o Specimen: Blood Film
o Dye: Diazo (alkaline pH)
o Substrate: Naphthol-phosphate
o Product: insoluble blue compound
Alkaline phosphatase works on the substrate
Enzyme + substrate = product (insoluble blue compound)
o Evaluate 100 neutrophils
▪ 0 no staining
▪ 1+ faint/diffuse
▪ 2+ pale/moderate
▪ 3+ strong blue ppt
▪ 4+ deep blue ppt without visible cytoplasm
o Reference interval: 15-170
o The mean score of two examiners is reported, and they should agree within 10%
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• BM transplantation has been successful in CML, and imatinib mesylate, a tyrosine kinase inhibitor, produces
remission in most cases.
NAP STAINING
Evaluate 100 neutrophils
0 x 93 = 0
1+ x 5 = 5
2+ x 2 = 4
3+ x 0 = 0
4+ x 0 = 0
9 NAP score = ↓
2. Polycythemia Vera
• characterized by excessive proliferation of erythroid, granulocytic and megakaryocytic elements in the bone
marrow (panmyelosis). Splenomegaly is usually present
• ↑ RBC, granulocytes, megakaryocytes, except for lymphocytes
• The increased red cell mass produces blood hyperviscosity, often resulting in cardiovascular disease
• Treatment includes:
o therapeutic phlebotomy (during early stages)
o alkylating agent hydroxyurea
o JAK inhibitors (e.g. ruxolitinib)
Polycythemia
1. Absolute – Hct & Red cell mass
a. Primary - ↑ RBC ct, ↓ EPO levels (ex: PV)
b. Secondary - ↑ RBC ct, ↑ EPO levels (ex: Renal tumors, hypoxic conditions:
smoking, ↑ altitude areas)
2. Relative – reduction in plasma volume
- ↑ Hct, Normal red cell mass (ex: Dehydration, burns, diuretics)
3. Essential Thrombocythemia
• clonal MPN with increased megakaryopoiesis and thrombocytosis
• Clinical manifestations
o vascular occlusions (thromboses in major arteries and veins)
o splenic infarcts resulting to atrophy
o pulmonary emboli and neurologic complications
o bleeding in mucous membranes
Normal platelet count: 150-400 x 109/L
• Lab findings:
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o Thrombocytosis (>400 x 109/L)
o Platelets often appear normal, but there could be presence of giant bizarre platelets, platelet aggregates,
micromegakaryocytes, and megakaryocyte fragments
o Leukocytosis is usually present and ranges from 22 to 40 x 109/L.
o Segmented neutrophils may be increased
• Treatment includes:
o Plateletpheresis
o Hydroxyurea therapy
o Low-dose aspirin
o JAK2 inhibitors
4. Primary Myelofibrosis
• It is the least common but most aggressive form of MPN
• Chronic, progressive clonal panmyelosis characterized by megakaryocytic and often granulocytic hyperplasia
with varying degrees of reactive fibrosis of the marrow & extramedullary hematopoiesis production of
blood cells outside the bone marrow
• Splenomegaly is present in 90% of patients with PMF, and 50% have hepatomegaly
• CBC findings:
o Normocytic anemia, anisocytosis, and poikilocytosis including Dacryocytes
o Pancytopenia resulting from fibrosis of BM
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o Dysmegakaryopoiesis (abnormal morphology)
▪ Giant platelets
▪ Abnormal granulation of platelets
▪ Abnormal nuclear shapes in the megakaryocytes
▪ Circulating micromegakaryocyte
• FAB Classification of MDS:
1. Refractory anemia
2. Refractory anemia with ring sideroblasts (RARS)
3. Refractory anemia with excess blasts (RAEB)
4. Chronic myelomonocytic leukemia
5. Refractory anemia with excess blasts in transformation (RAEB-t)
Lymphoproliferative disorders
• Includes the Lymphoid leukemia and Lymphoma
• classified by WHO thru their biological features and not the anatomical distribution
3. Burkitt Lymphoma
• aggressive cancer of mature B cells associated with a fulminant clinical presentation
• has three subtypes: endemic, sporadic, and HIV associated
• morphologic findings:
o medium-sized lymphoid cells which have deeply basophilic cytoplasm (strong blue); highly vacuolated
o Bone marrow and lymph node biopsies may show a classic “starry sky” appearance on Low power field
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5.
Diffuse Large B-cell Lymphoma
• Most common form (25-30% of cases) of Non-Hodgkin’s Lymphoma
• Clinical presentation is a rapidly expanding, painless lymphadenopathy in one or more sites
• Lab findings:
o Large lymphoid cells with diffuse histologic growth pattern.
o Surface markers: CD5, CD10, CD30
• Sezary syndrome
o Aggressive lymphoma with leukemic presentation (peripheral blood
involvement).
o characterized by erythroderma and generalized lymphadenopathy.
o An absolute Sézary cell count greater than 1 x 109/L is required to make a
diagnosis.
Hodgkin’s Lymphoma
• primarily a lymph node-based disease
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• has two distinct forms which differ pathologically and clinically:
o Classic HL
▪ makes up 95% of cases
▪ characterized by the presence of REED-STERNBERG CELLS
“owl’s eye” appearance
has abundant cytoplasm & bilobed nucleus with prominent eosinophilic nucleoli
B-cell origin and almost never found in peripheral blood (lymph node biopsy)
Surface markers: CD15 and CD30
o Lymphocyte-predominant HL
▪ malignant cell is a lymphocytic histiocytic cell
L&H cells are large with scant cytoplasm and a folded single nucleus;
they are labeled “popcorn” cells because of their distinct morphology.
50% express epithelial membrane antigen (EMA)
Specimen collection
• K2 EDTA less shrinkage of RBC; is the most common anticoagulant for routine hematology testing (CBC)
• Blood for CBC should be analyzed within 6 hrs (if stored at room temp) or within 24 hrs (4°C)
o Consequences: old specimen – RBC swelling, ↑MCV = presence of vacuoles on WBCs,
disintegration of platelets
• Peripheral blood films should be prepared within 3-4 hrs of collection
o Each of the four corner Large Squares is subdivided into 16 small squares
o Center Large Square is subdivided into 25 small squares
o Distance between each counting surface and the coverslip is 0.1 mm
Manual Cell Count
WBCs are counted in the four large corner squares
RBCs are counted in the central square but, 5 small square are used; 5 out of 25 small squares
are used for RBC count
Platelets are counted in whole area of central large square; 25 out of 25 small squares
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• General Formula
Cells counted x Dilution Factor
Area counted x 0.1mm
AREA
1 WBC square = 1 mm2
1 RBC square = 0.04 mm2
Platelet - Central large square = 1 mm2
Sample computations:
1. WBC count
145 WBCs are counted on four WBC large squares.
Dilution used by the technologist is 1:20
Formula: Cells counted x Dilution Factor
Area counted x 0.1mm constant (depth)
145 x 20 = 2,900
4 mm2 x 0.1 mm2 0.4 mm2
= 7,250/mm2 = ML = x 10 6/L
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= 7.25 x 10 9/L
Si unit = x 10 9/L
2. Platelet count
3. RBC count
Hemoglobin Determination
1. Visual Colorimetry
• Acid Hematin (Sahli-Hellige method)
o Procedure: 20 uL blood + 0.1N HCl = Acid hematin
o Allow the mixture to stand for _________ to develop color
o Mixture is then diluted using distilled water until its color matches the brown glass standard
o Hb F, Hi, HbCO and SHb cannot be converted into acid hematin
• Alkali Hematin
o Procedure: 20 uL of blood + 0.1N NaOH = Alkali hematin
o Hb F is resistant to akali denaturation
2. Photoelectric Colorimetry
• Cyanmethemoglobin method (HiCN)
o Recommended because it measures all forms of Hb except Sulfhemoglobin
o Specimen: 20 uL blood
o Reagent: 5 mL Drabkin’s solution
Potassium ferricyanide
Potassium cyanide
Dihydrogen potassium phosphate
• Sources of error:
o Cyanmethemoglobin reagent is sensitive to light. It should be stored in brown bottle / dark area.
o Turbidity of sample can cause false-increase values
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High WBC count (>20 x 109/L) High Centrifuge reagent-sample mixture and measure the supernatant.
PLT count (>700 x 109/L)
Lipemia Mix 0.01 mL of patient’s plasma + 5 mL of cyanmethemoglobin reagent,
and use it as reagent blank
Hb S and Hb C Prepare 1:2 dilution with distilled water and multiply the result by 2
3. Indirect method
• Chemical (Principle: 1 gHb contains 3.47 g Fe)
• Gasometric (Principle: 1 g Hb contains 1.34 mL O2)
4. Gravimetric
• Specific gravity (CuSO4 method; sp. Gr. Of 1.053)
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Microhematocrit determination
• Ratio of volume of RBCs to that of whole blood expressed as percentage or in ratios
• Unreliable as an estimate of anemia
• Rule of Three
o Can be used to quickly check the results of Hgb and Hct
o Applies only to specimens that have normocytic normochromic RBCs
o Hgb x 3 = Hct (± 3)
Reticulocyte count
• used to assess the erythropoietic activity of the bone marrow
• A sample from EDTA-whole blood is stained with a supravital stain, such as new methylene blue
• Procedure:
o Mix equal amounts of blood and NMB stain (2 to 3 drops, or approximately 50 uL each)
o Allow to incubate at room temp for 3 to 10 minutes. Remix and prepare two wedge films.
o Observe under OIO and calculate % reticulocytes
o Formula:
o To improve accuracy, another tech counts the other film; counts should agree within 20%
o Inserting a Miller disc into the eyepiece of microscope can facilitate counting
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RBCs are counted in the smaller square, and retics are counted in the larger square
A minimum of 112 RBCs should be counted in the small square
o Formula:
o Decreased in:
Polycythemia / Pancytosis
Poikilocytes (Sickle cells, Spherocytes, Acanthocytes)
Microcytosis
Hypofibrinogenemia
Hypogammaglobulinema
Newborn
• Mechanical / Technical factors
• Temperature: 20-25°C
• Should be performed within 4 hours after blood collection if kept at room temp
• Anticoagulant: _____________________
• ESR columns:
o Westergren Method
Length = 300 mm
Ext bore diameter = 5.5 mm
Int bore diameter = 2.5 mm
Graduation = 0-200 mm
o Wintrobe Method
Length = 115 mm
Int bore diameter = 3 mm
Graduation = 0-100 mm
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Peripheral Blood Films
• Should be made within 3-4 hrs after blood is collected using EDTA o Possible problems that can
be encountered in certain conditions:
▪ EDTA-inducted platelet satellitosis
▪ EDTA-induced platelet clumping
• Types of blood films:
o Wedge method
▪ most convenient and most commonly used technique
▪ requires at least two glass slides (dimensions: 75 x 25 mm or 3 inch x 1 inch)
• HPO (40x)
o use to select the correct area of the film in which to begin the differential count
o WBC estimation can be performed (Factor used: ___________)
• OIO (100x)
o WBC differential count is performed
“Battlement” track pattern is preferred to minimize distribution
errors.
100 WBCs are counted and classified through the use of push-
down counters.
To increase accuracy, it is advisable to count at least 200 cells
when WBC > 40 x 109/L.
o Evaluation of RBC, WBC, and platelet morphology.
o Platelet estimation can be performed (Factor used: ___________)
REFERENCE RANGES
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BONE MARROW EXAMINATION
• Indications: Neoplasia diagnosis, BM failure, infections, monitoring of treatment •
• Before BM collection (< 24 hrs), blood is collected for CBC with blood film examination
• Site of collection:
o Preferred: __________________ of the pelvis.
o Others sites: Sternum, anterior medial surface of the tibia
• Types of needles that are used for collection:
o Jamshidi biopsy needle
o Westerman-Jensen biopsy needle
o University of Illinois aspiration needle
o Snarecoil biopsy needle
• Preparation of BM samples.
o Direct aspirate smears
o Anticoagulated aspirate smears
o Crush smears of aspirate
o Buffy coat smears
o Core Biopsy Imprints / Touch Preparations
o Histologic Sections / Cell Block of the Core Biopsy
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Other Definitive Tests on BM samples
• Cytochemical studies
• Cytogenetic studies
• Molecular studies
• Flow cytometry
• Fluorescence in situ hybridization
AUTOMATION
• Electronic impedance
o based on the detection and measurement of changes in electrical resistance produced by cells as
they traverse a small aperture.
o electrical resistance between the two electrodes, or impedance in the current, occurs as the cells
pass through the sensing aperture, causing voltage pulses that are measurable.
The number of voltage pulses is proportional to ______________________.
The height of the voltage pulse is directly proportional to _________________.
o Blood sample is distributed into two channels: RBC/PLT and WBC/HGB.
o The data are plotted on a frequency distribution graph, or volume distribution histogram.
o The histogram produced depicts the volume distribution of the cells counted.
• Radiofrequency
o usually used in conjunction with electronic impedance
o the cell interior density is proportional to pulse height or change in the RF signal.
o Conductivity is determined using a high frequency electromagnetic probe that provides
information on the cell internal constituents.
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CAUSES OF ERRONEOUS RESULTS WITH AUTOMATION
Cold agglutinins
Microcytes or schistocytes
Platelet clumps
Old specimen
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