Introduction and Blood

Question 1

“Formed elements”

Answer 1

Non-plasma component of blood

Includes nucleated cells, erythrocytes, and platelets

Question 2

Resolution of a standard light microscope

Answer 2

0.2 μm

Question 3

Haemotoxylin

Answer 3

Hematoxylin (the blue dye) is a basic dye which will stain acidic structures a bluish color. Structures that contain DNA or RNA have affinity for this dye (i.e. nucleus), as DNA and RNA are acidic. Structures that react with hematoxylin are referred to as basophilic (base loving), as they bind to the basic dye.

Question 4

Eosin

Answer 4

Eosin (the pink dye) is an acidic dye that will stain basic components of the cell pink. Since cytoplasmic proteins are typically basic, the cytoplasm is usually stained pink. Components of the cell that react with eosin are referred to as acidophilic or eosinophilic.

Question 5

Wright-Giemsa

Answer 5

The Wright-Giemsa stain (sometimes just referred to as Giemsa) is frequently used to stain individual cells (as opposed to tissues). It is particularly helpful for staining blood (most of the images you will see in the reading specific to the blood session that accompanies this are actually Wright-Giemsa stains, and not H&E stains) and bone marrow. However, the general “nucleus blue, cytoplasm pink” rule still applies

Question 6

Trichrome Stain

Answer 6

The Trichrome stain is a variant of H&E that adds a third dye that specifically stains collagen a blue-green color, and is useful for visualizing connective tissue. A similar stain can be used to visualize elastin fibers in elastic connective tissue.

Question 7

transmission electron microscopy

Answer 7

The electron beam passes through the section. This method is useful for visualizing cells at the ultrastructural level (i.e., organelles and other subcellular structures below the limit of resolution of light microscopy).

Question 8

scanning electron microscopy

Answer 8

Uses a moving electron source to “scan” the surface of the specimen, which is coated with an electron dense material (gold is commonly used). The electrons bounce off of the surface of the sample, and will be collected by a detection system, which generates a 3D image of the sample being studied. This technique is useful for studying whole cells and tissues

Question 9

Relative sizes of bacteria, erythrocytes, and megakaryocytes.

Answer 9

Question 10

Relative size of erythrocyte and virus

Answer 10

Question 11

Relative size of virus and protein

Answer 11

Question 12

Myeloperoxidase staining

Answer 12

Question 13

Immunohistochemistry

Answer 13

In immunohistochemistry, a section of tissue is placed on a slide, and an enzyme-linked antibody is placed on the tissue. The antibody will bind to its specific antigen. Excess, unbound antibody is washed away. The position of the antibody is then visualized using a substrate for the antibody that generates a visible product.

It is sometimes called immunoperoxidase staining (“impox” for short), because the enzyme linked to the antibody is usually horseradish peroxidase.

Question 14

Immunofluorescence

Answer 14

very similar in concept to immunohistochemistry, but the antibody is linked to a fluorescent molecule rather than an enzyme, and is activated when it is exposed to light of a particular wavelength

Question 15

in situ hybridization

Answer 15

We take advantage of the principle of sequence complementarity: nucleic acid “probes” with sequence complementary to the region of interest are used to identify a particular DNA or RNA sequence target.

The probe is typically labeled with a visible signal (fluorophore or enzyme) that allows it to be detected. When the probe is fluorescent, the method is referred to as fluorescence in situ hybridization, or FISH.

Question 16

next-generation sequencing

Answer 16

instead of sequencing one long sequence, many short pieces of DNA (“reads”) are sequenced in parallel on a chip, then tiled together to get the full sequence. This allows sequencing to be performed much more cheaply and efficiently than using older methods.

“shotgun”

Question 17

Plasma

Answer 17

Unclotted liquid phase of blood

Question 18

Serum

Answer 18

post-clot liquid phase of blood

lacks fibrinogen and clotting factors

Question 19

Albumin

Answer 19

The most abundant plasma protein (it makes up 55% of circulating protein all by itself). It has a vital role in maintaining plasma oncotic pressure – without it, fluid would leak into the tissue.

Question 20

Gamma globulin

Answer 20

antibodies (all, not just IgGs)

Question 21

Blood electrolyte concentrations

Answer 21

Question 22

Thrombocyte

Answer 22

another name for platelet

Question 23

Number of erythrocytes in 1 microliter of adult blood

Answer 23

5x10⁶ per µl

Question 24

“central pallor”

Answer 24

Indent in erythrocytes

Question 25

Erythrocyte half-life

Answer 25

half-life in circulation of ~120 days

Question 26

hemostasis

Answer 26

blood clotting

Question 27

Answer 27

A Neutrophil

B Eosinophil

C Basophil

D Monocyte

E T Lymphocyte

F Plasma Cell

Question 28

Neutrophils

Answer 28

The most abundant of the leukocytes and usually comprise 50-70% of white blood cells (even higher during an acute inflammatory response). Neutrophils are the first responders to tissue injury and are the major mediators of the acute inflammatory response.

Question 29

Eosinophils

Answer 29

comprise up to 3% of circulating leukocytes in a normal adult. They are about 15 μm in diameter, same as neutrophils, but can be distinguished by their bilobed nuclei and bright pink (eosinophilic) granules. Eosinophils function in defense against helminthic infections (parasitic worms), and play an important role in the pathology of allergy.

Question 30

Basophils

Answer 30

The least numerous of the leukocytes, comprising about 0.5% (or roughly 1 in 200 cells). They are about the same size as neutrophils and eosinophils, but distinguished by their dark blue (basophilic) granules that typically obscure the nucleus from view.

Basophils release acute inflammatory mediators, including histamine and serotonin, during acute hypersensitivity reactions. If you see more than one or two basophils on a blood smear, something is most likely wrong with the patient. In particular, basophilia (too many basophils – see note on terminology) should make you suspect a particular hematologic malignancy, chronic myelogenous leukemia (CML)

Question 31

Lymphocytes

Answer 31

Typically comprise 10-20% of circulating leukocytes and include B, T, and NK cells (these cannot be distinguished morphologically – you need to use methods such as flow cytometry that assess cell surface marker expression to tell them apart). They are 7-8 μm in diameter. As a general rule, the nucleus of a resting lymphocyte should be about the same size as a red cell.

A resting lymphocyte should have a round nucleus, condensed chromatin (why?), and very little cytoplasm (again, why?).

Question 32

Monocytes

Answer 32

10-20% of circulating leukocytes, similar to lymphocytes. They are the largest circulating cells in normal blood, about 25 μm in diameter

In addition to their size, they can be identified by their folded (often bean-shaped) nuclei and bluish-gray cytoplasm without granules. When attracted to tissue by molecular signals (chemokines) during chronic inflammation, monocytes migrate out of the blood and into tissue, where they become macrophages and play an important role in “cleaning up” tissue damage and killing pathogens.

Question 33

–cytosis

Answer 33

too many of a cell type

Question 34

–penia or -cytopenia

Answer 34

too few of a cell type

Question 35

Anemia

Answer 35

too few red cells

Question 36

–philia

Answer 36

often used in place of –cytosis to indicate too many of a cell type.

Question 37

hematocrit

Answer 37

Historically, red cell mass was assessed using the hematocrit. To measure hematocrit, whole blood is placed in a very thin glass tube and spun down in a centrifuge. The ratio of the height of the red cell pellet to the total height of the column is the hematocrit (normal is around 36-45%; higher in men, lower in women). This method is quick, cheap, and easy – all you need is a tube, a centrifuge, and a ruler – and is still used in resource-poor settings

Most hospital labs that use automated instrumentation instead measure the concentration of hemoglobin in lysed blood as a surrogate for red cell mass (normal is 12-15 g/dl). The hematocrit is then calculated based on the hemoglobin concentration.

Question 38

hematopoiesis

Answer 38

The process of formation of the formed elements in the blood

Nearly all hematopoiesis in adults occurs in the bone marrow. (Interestingly, it occurs in the spleen and liver in the fetus, and migrates to the marrow during the last few months of gestation.) The bone marrow contains the precursors of all the formed elements of the blood: myeloid precursors (give rise to granulocytes, lymphocytes, and monocytes), erythroid precursors, and megakaryocytes, the precursors of platelets

Question 39

Turnover of erythrocytes, platelets, neutrophils

Answer 39

Question 40

dynamic flexibility of bone marrow

Answer 40

the ability to increase production of blood cells in response to stress

Question 41

hematopoietic stem cell differentiation pathways

Answer 41

Question 42

“Left Shift”

Answer 42

When blood progenitor cells exit the blood marrow early in response to stress and finish proliferating and differentiating in the bloodstream

Question 43

erythropoietin

Answer 43

Production of erythrocytes, or erythropoiesis, is regulated by the hormone erythropoietin

EPO is a small (166 amino acid) glycoprotein hormone produced by specialized cells in the kidney (the juxtaglomerular cells)

Production of EPO (and by extension, of erythrocytes) occurs in response to hypoxia, which triggers a signaling cascade involving the transcription factor HIF1α. Once in the nucleus, HIF1α drives transcription of EPO

Question 44

Erythropoetin in action

Answer 44

Once released into the blood, EPO acts on the earliest erythrocyte precursors in the bone marrow (BFU-E and CFU-E; see diagram on page 19) to drive them to proliferate and prevent them from undergoing apoptosis, resulting in increased production of mature erythrocytes. Once the red cell mass increases, and the hypoxic state is resolved, then HIF1α will be inactivated, and EPO production will decline. This is a classic example of a negative feedback loop.

Question 45

thrombopoietin

Answer 45

Production of platelets, or thrombocytes, is regulated by the hormone thrombopoietin (TPO).

TPO is produced at a relatively constant rate by the liver.

Both megakaryocytes (platelet precursors) and platelets themselves express the receptor for TPO. However, only megakaryocytes have the ability to respond to TPO and make platelets. The mature platelets therefore act as a “sink” for TPO. If there is a decline in the number of platelets, more TPO will bind to receptors on megakaryocytes, resulting in an increase in platelet production. These additional platelets will restore the balance of TPO bound to platelets vs. megakaryocytes, and production will return to normal.

Question 46

Leukocyte hematopoiesis

Answer 46

Question 47

Mononuclear Cells

Answer 47

Lymphocyte, monocyte, or macrophage

Question 48

Quantitative erythrocyte, platelet, and leukocyte measurement by light scattering

Answer 48

Question 49

Spherocytosis

Answer 49

lack of erythrocyte pallor

spherical erythrocytes