MLAB 1315- Hematology
Keri Brophy-Martinez
Unit 2 : Hematopoiesis
Unit 3 : Hematopoietic
Organs
Chapter 2: Terms
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Homeostasis
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Maintenance of an adequate number of cells to
carry out the functions of the organism.
Hematopoiesis
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Dynamic formation and development of blood
cells, normally in the bone marrow
Process responsible for the replacement of
circulating blood cells.
Review of Cell Morphology
Cell Morphology
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Cell Membrane
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Outer boundary of the cell
Allows passage of nutrients, ions and
information between cytoplasm and the
exterior
Phospholipid bilayer
Cell Membrane: Phospholipid Bi-layer
Cell Morphology

Cytoplasm: Location of metabolic activities
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Golgi complex: involved in formation of gamma globulins,
lysosomes, specific granules and other cellular
components.
Ribosomes: RNA, synthesize proteins
Endoplasmic reticulum: Communication system with the
nucleus, smooth or rough
Mitochrondria: furnish cell with energy
Lysosomes: granules with hydrolytic enzymes.
Plasma membrane: 2 layers of phospholipids,
surrounding cytoplasm
Cell Morphology
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Nucleus:
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Chromatin
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Nucleolus/Nucleoli
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Pale staining patches rich in RNA
Parachromatin
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Dark staining chromosomal material composed of DNA and
proteins.
DNA regulates all cellular functions
Zones of pale staining material between chromatin clusters
Nuclear membrane

Double membrane which surrounds the nuclear contents.
Maturation Characteristics of
Blood Cells
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Cytoplasm
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Immature: stains deep blue due to abundant
RNA, scant amount of cytoplasm, no granules
More mature: blue color decreases as RNA
decreases, granulocytes acquire non-specific
granules, cytoplasm amount decreases
Mature - granules (if present) become more
specific
Maturation Characteristics of
Blood Cells
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Nucleus
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Immature - round or oval, very large occupying
most of cell, active DNA and RNA replication
(dividing), fine chromatin pattern, nucleoli
present
More mature - size decreases, chromatin (DNA)
becomes more coarse and clumped, # of
nucleoli decreases
Mature - nucleus assumes final shape, no
nucleoli, chromatin is clumped, no division, in
RBC’s nucleus has been extruded
Overall Changes in Cell
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N/C ratio: nuclear to cytoplasmic ratio
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Immature “blast” cell has a large nucleus, small amount of
cytoplasm. N/C ratio is high
N/C ratio decreases with maturity as the nucleus decreases in
size and the cytoplasm becomes more abundant.
Size
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Immature cells are larger than mature cells
The size of almost all cell lines decreases with maturity
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Exception is thrombocyte/ platelet precursor in the bone
marrow increase in size with maturation
Lab Method: Wright’s Stain
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Stain used to
identify cells on
blood smear
Eosin is acid stain stains red
Methylene is basic
stain - stains
shades of blue
Cell Growth Factors
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Cytokines
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Growth factors secreted by cells for the purpose of cell-to-cell
communication. They stimulate the multipotential stem cells to
proliferate and differentiate.

Types
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Colony stimulating factors (CSF): These regulate blood cell development.
CSF’s are being synthetically manufactured to treat diseases.

G-CSF: Stimulate granulocyte production, Used to treat cancer

and AIDS patients with low WBC. Trade name is Neupogen
GM-CSF - Stimulate granulocyte-macrophage production,
Used to treat cancer patients with low WBC. Trade name is Leukine.

EPO - Stimulate erythrocyte production, Used to treat chronic
anemia caused by renal failure, increase RBC prior to surgery which
may cause blood loss, illegally by athletes to boost performance.
Trade name is Epogen or Procrit.
Cell Growth Factors
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Interleukins
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Wound healing, activating lymphocytes,
assisting in the growth of transplanted or
damaged bone marrow.
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Cluster Designation (CD)
nomenclature
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Using monoclonal antibodies and flow cytometry techniques, it is
possible to identify specific populations of hematopoietic cells.
Hematopoietic Organs
Hematopoiesis: blood formation
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Intramedullary
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Origin of blood cells and sequential sites of
normal blood production within the bone marrow
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Fetus
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Yolk sac
Liver and spleen
Bone marrow (all bones)
Adult
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Child up to teen years - all bones
18 years and up - flat bones (sternum, ribs, pelvis,
vertebra, skull)
Adults- bone marrow
Hematopoiesis
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Extramedullary hematopoiesis
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Blood is produced in the spleen, liver and other
tissues when demand is increased.
Storage Pools
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Stem cell pool
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Multipotential stem cell
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Unipotential (committed) stem cell
Bone marrow pool
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Granulocytes - proliferation & maturation/storage
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Thrombocytes - proliferation & maturation
Erythrocytes - proliferation & maturation
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Peripheral blood pool
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Granulocytes - 50% storage, 50% functional
Thrombocytes - 30% storage, 70% functional
Erythrocytes - 100% functional
Bone Marrow
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Bone marrow differentiates into
myeloid, erythroid and lymphoid cell
lineages under the influence of
cytokines or growth factors.
Organized around bone vasculature
Function: supply mature hematopoietic
cells into peripheral blood and respond
to demands
Indication for Bone Marrow Studies
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Hematologic diseases
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Anemia, erythrocytosis, polycythemia
Leukopenia and unexplained leukocytosis
Appearance of abnormal or immature cells in the peripheral
blood circulation
Thrombocytopenia or thrombocytosis
Systemic disease
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Solid malignant tumors elsewhere in the body - done at initial
diagnosis to determine the degree of tumor spread and to stage
the malignancy
Infections known as “fever of unknown origin” (FUO) in which
organisms may be within the marrow.
Certain histiocytic diseases
Procedure: Bone marrow
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Bones which have active hematopoiesis in
adults in order of preference for performing
bone marrow procedure:
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Posterior superior iliac crest - biopsy and
aspiration almost always done at this site
Sternum - used for aspiration only
Anterior superior iliac crest
Spinal processes
Anterior tibia used for infants and small children
Procedure
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Needles used for procedure
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Biopsy - Jamshidi trephine needle (8 or
11 gauge)
Aspiration - University of Illinois (U of I)
needle
Laboratorian’s role in bone marrow
procedure
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Provide the bedside equipment necessary for the procedure. Be
sure you know what tests the physician wants so you have the proper
specimen containers and all necessary equipment. Sometimes the
physician will perform bilateral biopsies for disease staging.
Practice sterile procedure in handing the physician items needed for
the aspiration and biopsy.
Receive the aspirate syringe from the physician and quickly prepare
smears at bedside and place in appropriate anticoagulated tubes
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Direct smear straight from syringe
Crush prep from marrow spicules
Receive other syringes of marrow if tests are requested such as
Leukemia/Lymphoma Profile (LLP or Flow Cytometry),
Immunophenotyping (chromosome studies), cultures (routine, viral or
fungal). Transfer into appropriate tubes.
Receive the biopsy specimen from the physician, prepare trephine
imprints and place in fixative
Processing the specimen in the
lab
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Place EDTA specimen (liquid aspirate) in Wintrobe tube and centrifuge at low
speed.
Measure layers formed by centrifugation
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FPV - fat and perivascular (used for iron stain)
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Plasma
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Buffy coat (myeloid:erythroid cells- M:E)
Normal is 4:1.
M:E is the ratio between all granulocytes and their precursors
and all nucleated red cell precursors.

RBC’s
Prepare and stain ME smears. Perform special stains if requested (further
discussion later).
Deliver clot remaining in syringe and biopsy to histology for processing.
Deliver other specimens obtained such as viral, fungal or routine culture
specimens to microbiology.
Complete paperwork and package specimens to be sent out to reference lab.
Information derived from
specimens
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Direct smear from syringe tip - evaluation of cellular morphology with
Wright’s stain
Particle (crush) smear - evaluation of cellularity and the relationship
of cells to each other
M:E smear - evaluation of hematopoietic cells and M:E ratio
FPV smear - evaluation of iron (use Prussian blue stain for iron)
Biopsy
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If marrow cannot be aspirated (“dry tap”), this is the only specimen for
examination
Examination for malignancy for clinical staging of lymphomas and
cancers
Examination of the architecture of the bone marrow and the cells in
their natural relationship to each other
Trephine imprint (touch prep) - examination of cells with Wright’s stain;
may be the only source to study cellular detail if an aspirate is not
obtained