Reading: RR&K, Chapter 9
SLIDE 17b blood smear, human (Wright)
(RR&K p211, Fig.1,2,3,4,5,6; p213, Fig. 1,2,3,4,5,6,7)
Scan the smear using the 40X objective lens. Look for an area where the smear is even and well stained. In a good area, the erythrocytes will be barely touching one another and will not be aggregated into rows of rouleaux. They will look circular in shape and, because they are biconcave, will appear more darkly stained on their periphery. Usually erythrocytes will appear pink; if they are bright red, all cells are overstained with acid dye; if they are grey, then all cells are over stained with basic dye. Use the oil immersion lens and, with the aid of your textbook, identify the cells listed in the check list.
Note: it may be difficult to find a basophil in your slide. Why?
Check list for SLIDE 17b:
SLIDE 17d red bone marrow smear, human (Giemsa)
(RR&K p202, Fig.9.11)
The different cell types in bone marrow can be most easily identified in smears. Bone marrow is aspirated from one of the bones containing red bone marrow (usually sternum or iliac crest in the adult). A drop of bone marrow is placed on a slide and then smeared across the slide. To study the smear with the microscope, look for a thin, evenly stained area. It is essential to find a good area on the smear. Most of it will be too thick or heavily stained. The RBC's on the smear establish the "eosinophilic" color. With the aid of an atlas and the table on the following page, identify the various cell types in both the erythrocytic series and the granulocytic series. In practice, it will not always be possible to distinguish one cell type from a cell that is slightly earlier or later in the lineage. This is because there is a range of characteristics for a given cell type and because cells present themselves variably as they are flattened and dried. Thus, neither you nor your instructor will be able to identify every cell on your slide. Nevertheless, the following is a series of steps to assist you in your identification:
First, look at the shape of nucleus and note whether there is a nucleolus present. A round nucleus with a nucleolus is either a proerythroblast or promyelocyte. These cells can be distinguished by the fact that promyelocytes contain red azurophilic granules. Round nuclei without a nucleolus will more than likely belong to an erythrocyte precursor.
In the erythrocyte series, look at the distribution of nuclear euchromatin and heterochromatin and the staining properties of the cytoplasm. During differentiation, the amount of active euchromatin decreases and the cytoplasm changes from basophilic to eosinophilic.
In the granulocyte series, look at the shape of the nucleus and the presence of azurophilic and specific granules in the cytoplasm. The shape of the nucleus will assist you in differentiating between myelocyte, metamyelocyte, band, and mature granulocytes. The specific granules will either be unstained, eosinophilic, or basophilic.
Check list for SLIDE 17d: 
| Nomenclature and Recognizable Developmental Stages of the Erythrocytic Series Table for SLIDE 17d |
|
|---|---|
| Nomenclature | Important features |
| Proerythroblast | Cells are round cells 13-18 um in diameter; large spherical nucleus with 1-2 distinct nucleoli; thin rim of slight to moderate basophilic cytoplasm (basophilia attributable to increasing numbers of free ribosomes; rER absent). These cells divide giving rise to: |
| Basophilic erythroblasts | Cells in this series divide and are slightly smaller than the proerythroblasts and have deeply basophilic cytoplasm. EM reveals considerable numbers of polyribosomes present (beginning of hemoglobin synthesis at this stage). Division of basophilic erythroblasts gives rise to: |
| Polychromatophilic erythroblasts | Cells have cytoplasm that is a dull gray to gray-green color due to a combination of basophilic staining (polyribosomes) and eosinophilic staining (hemoglobin). Continued division and maturation gives rise to: |
| Normoblasts (acidophilic or orthochromatic erythroblasts) | Cells have cytoplasm distinctly eosinophilic due to the accumulation of hemoglobin, but will exhibit a light blue or green coloration due to the basophilic nature of the ribosomes still present. The nucleus is small, dense, and eccentrically located. Extrusion of nucleus gives rise to: |
| Polychromatophilic erythrocyte (reticulocyte) | Cells are enucleated with a cytoplasm that is slightly more basophilic that the erythrocyte. |
| Erythrocyte | Cells are the mature red blood cells found in the circulation. |
| Nomenclature and Recognizable Developmental Stages of the Granulocyte Series Table for SLIDE 17d |
|
|---|---|
| Nomenclature | Important features |
| Promyelocyte | Cells are 16 um in diameter, slightly indented nucleus with distinct nucleolus, characterized by cytoplasmic azurophilic granules. During maturation there is an increase in number of granules and increase in cell size (24 um). As a result of one or more divisions late promyelocytes are produced. These are smaller in size. |
| Myelocytes | Cells are recognized by the appearance of secondary or specific granules. There is also a reduction in number of the azurophilic granules. These cells are further distinguished according to the type granules that develop, i.e., neutrophilic myelocyte (60%), eosinophilic myelocyte, basophilic myelocyte (3%). The nucleus of these cells is slightly indented. Cells in this series divide and give rise to: |
| Metamyelocytes (neutrophilic, basophilic and eosinophilic) | Cells show varying degrees of indentation, and elongation of the nucleus indicative of differentiation into the more mature juvenile or band forms. |
| Band (neutrophilic) | Cells are identified by their elongated nucleus that appears either U-shaped or S-shaped. |
| Mature (neutrophilic, basophilic and eosinophilic) | Cells are the mature forms found in the circulation. They have a multilobed segmented nucleus. |
SLIDE 07a neck, mouse (H&E)
(RR&K p208, Fig.9.15)
This section shows small patches of bone marrow with excellent preservation in the marrow cavities of the bones. Study the overall relationship of myeloid cells, fat cells and bone trabeculae. Identify sinusoids with their reticular (adventitial) cells, and sinusoidal lining cells (endothelial cells). Observe as many of the granulocytic and erythrocytic series as you can. Also note that cells of one lineage tend to occur in groups (noticeable especially with cells of the erythrocytic and neutrophilic series). Why?
Check list for SLIDE 07a:
SLIDE17p blood smear, leukemia (Wright)
(RR&K p211, Fig.1,2,3,4,5,6; p213, Fig. 1,2,3,4,5,6,7)
Leukemia is a malignant neoplasm of the hematopoietic stem cells arising in the bone marrow. It may cause an increase in the number of peripheral WBCs (leukemic leukemia > 15,000 WBC/mm3) or the number of WBCs may not be significantly increased (aleukemic leukemia < 15,000 WBC/mm3). Leukemia is classified on the basis of the cell type involved (myeloid versus lymphoid) and on the state of cell maturity. Acute leukemias are characterized by the presence of very immature cells (called blasts) and have a rapid, fatal course in untreated patients. Chronic leukemias are initially associated with well defined (mature) white blood cells. This blood smear (SLIDE 17p) comes from a 60-years-old male diagnosed in a routine physical examination with chronic granulocytic (myeloid) leukemia (CGL). This type of leukemia has a slow onset and usually occurs in older patients who complain of weakness, easy fatigability, malaise, weight loss and anorexia. Peripheral blood contains about 50,000 WBC/mm3, predominately mature forms of granulocytes. Identify and classify them. Promyelocyte, myelocytes and metamyelocytes may also be present. Try to identify these cells on your slide. Do you see any abnormality in the red blood cells?
Check list for SLIDE 17p:
Blood is connective tissue that is essential for: 1) delivering nutrients and oxygen, 2) removing cellular waste, 3) delivering hormones, 4) transporting elements of the immune system (antibodies, leukocytes, etc.). This unit examines both mature and precursor cellular elements of the blood.