Hematology Flashcards
What is the composition of whole blood?
RBCs, WBCs, Platelets, and Plasma
After centrifugation of whole blood, what does the buffy coat consist of?
WBCs and Platelets
What is the difference between plasma and serum?
Plasma - liquid portion of UNCLOTTED blood
Serum - liquid portion of CLOTTED blood
Does serum contain all coagulation proteins?
No, it lacks the fibrinogen group
Does plasma contain all coagulation proteins?
Yes
Describe the appearance of plasma.
hazy and pale yellow
Describe the appearance of serum.
clear and straw colored
What is the formula for calculating MCV?
What is the reference range for MCV?
80-100 fL
What does MCV measure?
the average/mean volume of RBCs (and size of RBCs)
What conditions is MCV INCREASED in?
- megaloblastic anemia
- hemolytic anemia with reticulocytosis
- liver disease
Note: normal in newborn.
macrocytic anemias
What conditions is MCV DECREASED in?
- iron deficiency anemia
- thalassemia
- sideroblastic anemia
- lead poisoning
microcytic anemias
Is MCV calculated or directly measured by automation?
directly measured by automation
What is the formula for MCH?
What is the reference range for MCH?
27-31 pg (pictograms)
What does MCH measure?
the average weight of hemoglobin in individual RBCs
What conditions is MCH INCREASED in?
macrocytic anemias
What conditions is MCH DECREASED in?
microcytic anemias
What RBC indices parallel MCV?
MCH
What is the formula for calculating MCHC?
What is the reference range for MCHC?
32-36% or g/dL
What is MCHC a measure of?
the average concentration of hemoglobin
What conditions is MCHC INCREASED in?
- presence of spherocytes
- error in RBC or HGB measurement
What conditions is MCHC DECREASED in?
- iron deficiency
- thalassemia
hypochromic RBCs
What is the reference range for RDW?
11.5-14.5%
What is RDW determined by?
the RBC histogram
What is the RDW increased in proportion to?
the degree of anisocytosis (variation in size)
What conditions is a high RDW seen in?
- post-transfusion
- post-treatment (e.g., B12, folic acid, iron therapy)
- idiopathic sideroblastic anemia
- in presence of 2 concurrent deficiencies (iron and folic acid deficiency)
What does the RDW help to identify?
anisocytosis
What is the normal hematocrit for a male?
40-54% (0.40-0.54 L/L)
What is the normal hematocrit for a female?
35-49% (0.35-0.49 L/L)
What id the normal hematocrit for a newborn?
48-68% (0.48-0.68 L/L)
Is the buffy coat included in the measurement of hematocrit?
no
What is the calculation for the hematocrit?
Hct (%) = (MCV x RBC)/10
What is PCV?
packed cell volume - another name for hematocrit
-it is the ratio of the volume of RBCs to the volume of whole blood
How do the spun hematocrit (manual) and the hematocrit derived from an automated cell counter differ?
-the automated hct is derived from the RBC and MCV, therefore it may be lower than the spun hematocrit in which there is always some trapping of plasma within the RBC column
-the difference is negligible unless there is a great deal of poikilocytosis and anisocytosis
What is the usual time and speed of centrifugation for a microhematocrit?
10,000 RPM for 5 minutes
How would a microhematocrit result be affected by centrifugation speeds and times that are lower than 10,000 RPM for 5 minutes?
-lower speeds or times leads to inadequate packing of the RBCs and a falsely elevated hematocrit
While performing a manual hematocrit, the technologist observes an increased buffy coat. What is the significance of this observation?
-the patient has an elevated WBC or extremely elevated platelet count
-the buffy coat should not be included in the measurement of the RBC column or the hematocrit will be falsely elevated
While reading a manual hematocrit a technician observes that the plasma is pink. How would this affect the hematocrit?
Hemolysis of the RBCs decreases the hematocrit.
A phlebotomist had a difficult time obtaining blood from a patient and was only able to fill the EDTA tube 1/4 full. How would this affect the hematocrit?
-automated hct - probably not affected
-microhematocrit - falsely decreased because the RBCs would be crenated by the excess anticoagulant
How does excess anticoagulant affect platelets?
it breaks them up which would cause a false increase in the platelet count
How does excess anticoagulant affect WBCs?
causes degenerative changes
A new phlebotomist had difficulty finding a vein in a patient and the tourniquet was left on for 3 minutes before the blood was obtained. How would this affect the hematocrit?
It would be elevated due to hemoconconcentration.
Which of the RBC indices is considered the most clinically useful?
MCV - it indicates the average size of the RBCs and is useful in the classification of anemias
Calculate the MCV, MCH, and MCHC for the following specimen. How should the RBCs appear on the blood smear?
RBC: 3.0 x 10^12/L
HGB: 6 g/dL
HCT: 20%
MCV = 67 fL
MCH = 20 pg
MCHC = 30 g/dL
The RBCs would appear microcytic (low MCV) and slightly hypochromic (MCHC).
What is the only clinical significance of a high MCHC?
hereditary spherocytosis
What is MCHC useful as an indicator of?
the presence of an interfering factor in the specimen (e.g., hemolysis, cold agglutinins)
A patient’s MCHC is reported as 37%. What abnormality should be looked for on the peripheral blood smear?
spherocytes
-if spherocytes are not present, one or more the RBC parameters may be incorrect
A patient’s MCHC is reported as 30%. What abnormality should be looked for on the peripheral blood smear?
hypochromic RBCs
-if hypochromic RBCs are not seen on the smear, one or more of the RBC parameters may be incorrect
What abnormality would be expected on the blood film when the MCV is high?
macrocytes
Explain how a blood sample can have a normal MCV when marked anisocytosis is observed on the blood smear.
MCV is an average - it can be normal when there is a combination of macrocytes and microcytes
-the RDW would be INCREASED
What abnormality would be expected on the blood film when the MCHC is low?
hypochromic RBCs
What is the “rule of three”?
QC check for RBC parameters of the CBC
What should one do if CBC values obtained do not follow the “rule of three”?
- check for abnormal RBC morphology on the blood smear
- check for specimen problem (e.g., lipemia, cold agglutinins)
- check for instrument error
What is the adult male reference range for hemoglobin?
14-18 g/dL (140-180 g/L)
What is the adult female reference range for hemoglobin?
12-15 g/dL (120-150 g/L)
A patient has a hemoglobin of 12.3 g/dL. What is the expected range for the patient’s hematocrit?
36.9 +/-3
What is the reference range for platelets?
150-450 x 10^3/uL (150-450 x 10^9/L)
What is the reference range for MPV (mean platelet volume)?
6.8-10.2 fL
What is the adult reference range for WBC count?
4.5-11.5 x 10^3/uL (4.5-11.5 x 10^9/L)
What is the adult male reference range for RBC count?
4.6-6.0 x 10^6/uL
(4.6-6.0 x 10^12/L)
What is the adult female reference range for RBC count?
4-5.4 x 10^6/uL
(4-5.4 x 10^12/L)
Relative count vs. Absolute count
-relative count - the amount of a cell type in relation to other blood components
-absolute count - the actual number of each cell type without respect to other blood components
What is relative lymphocytosis?
an increase in the % of lymphocytes (often associated with neutropenia)
What is relative polycythemia?
RBCS appear increased due to decreased plasma volume
What is absolute lymphocytosis?
a true increase in the number of lymphocytes
What is absolute polycythemia?
a true increase in red cell mass
What is a Romanowsky stain?
a nonvital (dead cell) polychrome stain
List the 4 Romanowsky stains.
GWML
- Giemsa stain
- Wright and Wright-Giemsa stain
- May-Grunwald stain
- Leishman stain
List the components of a Wright stain.
- methylene blue - a basic dye that stains acidic cellular components BLUE (DNA and RNA)
- eosin - an acidic dye that stains basic components red-orange (hemoglobin and eosinophilic cytoplasmic granules)
- methanol fixative - fixes cells to slide
- phosphate buffer (6.4)
What is Prussian blue?
a nonvital monochrome stain - stains specific cellular components
Lis the components of a Prussian blue stain.
- potassium ferrocyanide
- HCL
- safranin counterstain
What is Prussian blue used to visualize?
IHU
- iron granules in RBCs (siderotic iron granules)
- histiocytes
- urine epithelial cells
List the supravital (living cell) monochrome stains.
- new methylene blue
- neutral red with brilliant cresyl green as a counter stain
What is new methylene blue used for?
to precipitate RNA in reticulocytes => measure of bone marrow erythropoeisis
What is neutral red/brilliant cresyl green used for?
visualize Heinz bodies => G6PD deficiency and other unstable hemoglobin disorders
Are fixatives used in supravital stains?
no
Where does hematopoiesis occur in the fetus?
1-2 mo: yolk sac (primitive erythroblasts; embryonic hgb - Gower I, Gower II, & Portland)
3-6 mo: liver spleen (liver is primary site)
7 mo - 4 yr: bone marrow (ALL marrow is active)
List the organs of the reticuloendothelial system (RES).
- bone marrow
- spleen
- liver
- thymus
- lymph nodes
List the functions of the reticuloendothelial system.
- hematopoiesis
- phagocytosis
- immune defense
List the location of active marrow in adults.
flat bones
- skull
- sternum
- pelvis
- ribs
- vertebrae
Which hormone regulates the rate of erythropoiesis?
Erythropoietin (EPO)
Explain blood cell maturation.
Where and when is EPO produced?
-produced by the kidneys in response to cellular hypoxia and stimulates stem cells in the bone marrow, primarily CFU-E, to mature into erythrocytes
Which cells are derived from a common myeloid progenitor (CMP) cell in the bone marrow?
-erythrocytes
-granulocytes
-monocytes
-thrombocytes (platelets)
What is the precursor of the platelet?
megakaryocyte
-platelets are fragments of megakaryocyte cytoplasm
Where are lymphocytes produced?
- primary lymphoid tissue (thymus and bone marrow)
- spleen
- lymph nodes
- intestine-assoc. lymphoid tissue
- tonsils
What is the normal lifespan of an erythrocyte?
120 days
-phagocytic cells of the RES removed aged RBCs from circulation
-iron and globin chains are recycled
-heme is degraded and excreted as bilirubin
In a normal individual, what % of RBCs is replaced daily?
1%
-reticulocytes are released from the bone marrow into the circulation to replace aged cells removed by the RES
In the primary lymphoid tissue, what is the site of pre-B cell differentiation?
bone marrow
In the primary lymphoid tissue, what is the site of pre-T cell differentiation?
thymus
What is antigen-independent lymphopoiesis?
pre-B and pre-T cell differentiation in the primary lymphoid tissues
What is antigen-dependent lymphopoiesis?
lymphopoiesis that depends on antigenic stimulation of T and B lymphocytes
-occurs in secondary lymphoid tissue - lymph nodes, spleen, gut-associated tissue (Peyer’s patches)
What happens to the nuclear:cytoplasmic (N:C) ratio as most cells mature?
the ratio decreases as the volume of the nucleus decreases
What is the best indicator of the age of a cell?
that amount of chromatin clumping in the nucleus
What is the color of the cytoplasm in blasts?
royal blue due to the presence of RNA
What is the significance of nucleoli in a cell nucleus?
they are sites of ribosomal RNA synthesis
-seen in the nuclei of immature cells and reactive lymphocytes
-indicate that the cell is capable of mitosis
How are leukocytes classified?
- phagocytes (granulocytes, monocytes)
- immunocytes (lymphocytes, plasma cells, monocytes)
List the granulocytes.
- neutrophils
- basophils
- eosinophils
What leukocyte is the first to reach the tissues and phagocytize bacteria?
neutrophils
Which leukocyte arrives at the site of inflammation after neutrophils?
macrophages (monocytes in tissues)
What type of immunity do T lymphocytes provide?
cellular immunity
What lymphocyte makes up 80% of lymphocytes in the blood?
T lymphocytes
When activated, what to T lymphocytes produce?
cytokines/interleukins
What do B lymphocytes develop into in the tissue?
plasma cells
What do B lymphocytes produce?
antibodies
What type of immunity do B lymphocytes provide?
humoral immunity
What is the function of natural killer (NK) lymphocytes?
destroy tumor cells and cells infected with viruses
-also known as large granular lymphocytes (LGLs)
What is the function of eosinophils?
modulate allergic response caused by basophil degranulation
What is the function of basophils?
mediate immediate hypersensitivity reactions (type I, anaphylactic)
What are surface proteins expressed by specific cell lines at different maturation stages called?
CD markers
CD2, CD3
lymphoid, pan T cells
CD4
helper/inducer cells
CD8
suppressor/cytotoxic T cells
CD13
pan myeloid
CD11c, CD14
monocytes
CD19, CD20
lymphoid, pan B cells
CD33
pan myeloid cells
CD34
stem cell marker (lymphoid and myeloid precursor)
CD16, CD56
NK cells
Name the stages of development in the erythrocytic series, using 2 systems of nomenclature.
- rubriblast (pronormoblast)
- prorubricyte (basophilic normoblast)
- rubricyte (polychromatophilic normoblast)
- metarubricyte (orthochromic normoblast)
- diffusely basophilic erythrocyte (polychromatophilic erythrocyte/reticulocyte)
- mature erythrocyte
Pronormoblast
14-24 um
N:C ratio is 8:1
royal blue cytoplasm
fine chromatin
1-2 nucleoli
normally confined to the bone marrow
-14-24 um
-N:C ratio is 8:1
-royal blue cytoplasm
-fine chromatin
-1-2 nucleioli
-normally confined to the bone marrow
Pronormoblast
Basophilic normoblast
12-17 um
N:C ratio of 6:1
deep blue cytoplasm
chromatin is coarser with slightly visible parachromatin
nucleoli usually not visible
normally confined to the bone marrow
-12-17 um
-N:C ratio of 6:1
-deep blue cytoplasm
-chromatin is coarser with slightly visible parachromatin
-nucleoli usually not visible
-normally confined to the bone marrow
basophilic normoblast
polychromatophilic normoblast
10-15 um
N:C ratio is 4:1
cytoplasm is polychromatophilic due to hemoglobin production
chromatin is clumped with distinct areas of parachromatin (spoke-like pattern)
last stage to divide
normally confined to the bone marrow
-10-15 um
-N:C ratio is 4:1
-cytoplasm is polychromatophilic due to hemoglobin production
-chromatin is clumped with distinct areas of parachromatin (spoke-like pattern)
-last stage to divide
normally confined to the bone marrow
polychromatophilic normoblast
Orthochromic normoblast
-8-12 um
-N:C ratio is 1:2
-nucleus is pyknotic
-last nucleated stage
-normally confined to the bone marrow
-8-12 um
-N:C ratio is 1:2
-nucleus is pyknotic
-last nucleated stage
-normally confined to the bone marrow
orthochromic normoblast
polychromatophilic erythrocyte
-7-10 um
-no nucleus
-cytoplasm is diffusely basophilic (bluish tinge)
-reticulum seen with supravital stain
-0.5-1.5% of RBCs in adult peripheral blood
-7-10 um
-no nucleus
-cytoplasm is diffusely basophilic (bluish tinge)
-reticulum seen with supravital stain
-0.5-1.5% of RBCs in adult peripheral blood
polychromatophilic erythrocyte
mature erythrocyte
-7-8 um
-biconcave disk
-reddish-pink cytoplasm with area of central pallor 1/3 diameter of cell
-7-8 um
-biconcave disk
-reddish-pink cytoplasm with area of central pallor 1/3 diameter of cell
mature erythrocyte
What is the last nucleated stage in the development of an erythrocyte?
orthochromic normoblast (metarubricyte)
What is asynchronous erythropoiesis?
the nucleus and cytoplasm mature at different rates
What causes megaloblastic asynchronous erythropoiesis?
vitamin B12 and/or folic acid deficiency
Explain megaloblastic asynchronous erythropoiesis.
-nucleus lags behind cytoplasm in maturation
-cells grow larger without dividing
What characteristics of megaloblastic asynchronous erythropoiesis are seen on the peripheral smear?
oval macrocytes
What causes iron deficiency asynchronous erythropoeisis?
iron deficiency - reduces erythropoietin production
Explain iron deficiency asynchronous erythropoiesis.
-cytoplasm lags behind nucleus in maturation due to inadequate iron for hemoglobin synthesis
What characteristic is seen on the peripheral smear in iron deficiency asynchronous erythropoiesis?
microcytic, hypochromic RBCs
What is the best way to judge the size of an erythrocyte on a Wright-stained smear?
a normocytic RBC is approx. the same size, or slightly smaller than the nucleus of a mature lymphocyte
What is the first sign of accelerated erythropoiesis?
increased reticulocyte count
What diseases are macrocytes associated with?
- megaloblastic anemias (B12/folate deficiency)
- liver disease
- accelerated erythropoiesis
-normal in newborns
What diseases are associated with microcytes?
- iron deficiency anemia
- thalassemias
- sideroblastic anemia
- anemia of chronic disease
What is anisocytosis?
variation in RBC size - indicating a dimorphic population
What RBC parameter does anisocytosis correlate with?
RDW - esp. when it exceeds 15%
What diseases are associated with anisocytosis?
- post-transfusion
- post-treatment (B12, iron, folate)
- presence of 2 concurrent deficiencies (e.g., iron and B12)
- idiopathic sideroblastic anemia
What is poikilocytosis?
variation in the shape of RBCs
Echinocyte (Burr cell)
What is the clinical significance of echinocytes (burr cells)?
-membrane defect
- liver disease
- uremia
- hemolytic anemias
- TTP
- DIC
- carcinoma of stomach
- pyruvate kinase deficiency
What is the clinical significance of crenated RBCs?
-osmotic imbalance
-if seen in most cells in thin part of smear, do not report = probably artifact due to excess anticoagulant or slow drying
How can crenated RBCs be differentiated from burr cells?
Burr cells have more pointed projections as compared to the knobby projections of crenated cells.
-if the abnormality is seen in nearly all cells in the thin part of the smear, it is most likely artifact (crenation) due to slow drying of the smear or over-anticoagulation
-burr cells are seen in variable number throughout the smear
What is hypochromia?
the are of central pallor is greater than 1/3 the diameter of the RBC
spur cells (acanthocytes)
What is the clinical significance of acanthocytes (spur cells)?
-membrane defect (excessive cholesterol)
- alcoholic liver disease
- post-splenectomy
- abetalipoproteinemia
How are echinocytes and acanthocytes differentiated on a smear?
target cells (codocytes)
What causes target cells (codocytes)?
-excessive cholesterol in membrane
-hemoglobin distribution imbalance
What is the clinical significance of target cells (codocytes)?
- liver disease
- hemoglobinopathies (sickle cell, hemoglobin C)
- thalassemia
- iron-deficiency anemia
teardrops (dacrocytes)
What is the clinical significance of dacrocytes (tear drop)?
- anemias (megaloblastic)
- thalassemia
- extramedullary hematopoiesis (myelofibrosis)
sickle cells (drepanocytes)
schistocyte (RBC fragments)
What is the clinical significance of schistocytes?
-RBCs split by fibrin strands
- microangiopathic hemolytic anemias (DIC, TTP, HUS)
- prosthetic heart valves
- burns