Bone marrow evaluation is an important diagnostic tool for evaluating

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1 670 Vol. 24, No. 9 September 2002 CE Article #1 (1.5 contact hours) Refereed Peer Review KEY FACTS Bone marrow aspiration and core biopsy procedures are relatively simple, rapid, and inexpensive. The most frequent indications for performing a bone marrow examination include severe nonregenerative anemia, persistent neutropenia, nonregenerative thrombocytopenia, pancytopenia, atypical cells in blood, hypercalcemia, and hyperproteinemia. General categories of bone marrow disorders include hyperplasia, hypoplasia, aplasia, dysplasia, and neoplasia (e.g., leukemia). Comments? Questions? compendium@medimedia.com Web: VetLearn.com Fax: Collection and Assessment of Canine Bone Marrow University of Minnesota Douglas J. Weiss, DVM, PhD, DACVP Stephanie A. Smith, DVM, MS, DACVIM ABSTRACT: Bone marrow evaluation is an essential diagnostic tool for assessing bone marrow disorders that cannot be diagnosed by evaluating CBCs and other hematologic tests. The techniques for preparing bone marrow aspiration smears and core biopsies can be readily completed by practitioners. Initial cytologic evaluation of aspiration smears at the time of collection is useful for assessing the quality of the specimen and may reveal important preliminary information about the hematologic disorder. Bone marrow evaluation is an important diagnostic tool for evaluating hematologic disorders in dogs. 1 3 Collection of bone marrow specimens is relatively simple, rapid, and inexpensive and can be readily performed in clinical practice. 2,4 However, bone marrow evaluation depends on not only obtaining a sample but also preparing high-quality slides for cytologic evaluation and adequately preserving core biopsy specimens. Cytologic evaluation of bone marrow aspiration smears at the time of collection is important for determining whether marrow was obtained, assessing the cytologic quality of the specimen, and providing rapid initial information about the clinical condition. 2 The usefulness of the information obtained from bone marrow evaluation depends on whether there is a clear indication for performing the procedure, proper collection techniques, preparation of high-quality slides and core biopsy material, proper staining techniques, and interpretation of results with consideration for the case history, CBC, and other diagnostic information. INDICATIONS FOR BONE MARROW EVALUATION Appropriate use of bone marrow aspiration and core biopsy techniques depends on identifying a specific reason for performing the procedures. 2,3 In our experience, the most common reason that bone marrow evaluation fails to result in clinically useful information is that there is not a compelling reason to perform the procedure in the first place. Indications for bone marrow examination are usually derived from the case history, concurrent diseases, or changes in peripheral blood cell numbers or morphology. The most frequently identified hematologic indications for bone marrow examination include nonregenerative anemia, persistent neutropenia, nonregenerative thrombocytopenia, pancytopenia, atypical cells in the blood, unexplained hypercalcemia, hyperproteinemia/monoclonal gammopathy (i.e., multiple myeloma, lymphoma, ehrlichiosis, leishmaniasis, systemic fungal disease), staging of lymphosarcoma and

2 Compendium September 2002 Canine Bone Marrow Evaluation 671 mast cell tumor, and assessment of the feasibility of using chemotherapeutic drugs or other drugs known to damage bone marrow. 2,3 Alternatively, bone marrow examination is not usually indicated for most regenerative anemias, mild to moderate nonregenerative anemia associated with other disease conditions, regenerative thrombocytopenia (as determined by the presence of increased numbers of reticulated platelets in blood), and cytopenia associated with certain drug toxicities (e.g., sulfadiazine, quinidine, cephalosporins). 4,5 Figure 1 Illinois sternal biopsy needle placed in the iliac crest of a dog. Figure 2 Illinois sternal biopsy needle placed in the proximal femur of a dog through the trochanteric fossa. ASPIRATION BIOPSY TECHNIQUE Evaluation before performing bone marrow techniques should include a complete history and physical examination as well as a CBC on the day bone marrow is collected. 1 In general, a clotting factor abnormality or thrombocytopenia rarely results in significant hemorrhage during bone marrow collection; however, bleeding remains a potential problem. 2 If hemorrhage occurs, digital pressure applied to the biopsy site is usually all that is needed to prevent further blood loss. Several biopsy techniques have been described. 1 3 The most frequently used sites for bone marrow collection include the iliac crest, the trochanteric fossa of the proximal femur, and the proximal humerus. These sites are readily accessible and have greater hematopoietic activity than other sites. The choice of site depends on the experience and preference of the clinician. However, patient condition may influence site selection. Obese dogs have large fat layers over the ilium and trochanteric fossa, which make these sites less accessible. 2 The sites may not be palpable, and aspiration needle length may be too short to reach the sites. In these dogs, the proximal humerus is the most accessible site. In smallbreed dogs, aspiration of bone marrow from the proximal femur or humerus may result in lameness for several days. Lameness can be prevented by using the iliac crest. The site should be clipped and aseptically prepared. Sterile gloves should be worn, but a surgical drape is optional. Bone marrow aspirates and core biopsies can usually be performed with local anesthesia only or local anesthesia and mild sedation. 1 3 The skin, subcutaneous tissue, and periosteum must be anesthetized, and a small stab incision should be made to facilitate entry of the needle. Although a variety of needles are available, Illinois sternal needles (American Pharmaceal, Valencia, CA) are most frequently used for bone marrow aspirates, and Jamshidi needles (American Pharmaceal, Valencia, CA) are most frequently used for core biopsies. 3 Reusable (metal) and disposable (combination of metal and plastic) needles are available. Both needles have a stylet, which is essential for preventing plugging of the needle lumen with cortical bone. The Illinois sternal needle is 1 inch long, which is too short to reach the femur and ilium of larger dogs. To get additional length, the protective sleeve of the needle can be removed by unscrewing it. This provides a needle length of 2 inches, which is adequate for most large-breed dogs. If the ilium is used for aspiration, the entry point is the high point of the iliac crest (Figure 1). The dog can stand or be placed in dorsal or lateral recumbency. The needle is directed ventrally and slightly caudally into the crest. If the proximal femur is used, the dog should be placed in lateral recumbency and the greater trochanter located (Figure 2). The needle should be advanced parallel to the shaft of the femur and medial to the greater trochanter until it is firmly seated in the fossa. A good approach is to grasp and stabilize the stifle with the hand that is not manipulating the needle. If the humerus is used for marrow collection, the area of the greater humoral condyle should be tapped (Figure 3).

3 672 Small Animal/Exotics Compendium September 2002 Figure 3 Illinois sternal biopsy needle placed in the proximal humerus of a dog. Figure 4 A drop of bone marrow placed on a microscope, and a wedge-type smear that was made by touching a spreader slide to the drop of marrow. Note the presence of unit particles (i.e., white specks) in the drop and on the feathered edge of the smear. The humerus should be grasped and flexed and the needle inserted just medial to the greater condyle. Regardless of the site used, the subsequent procedures are the same. Once contact is made with the bone, the needle should be advanced with a firm, rotating motion. When the needle is firmly seated in the bone, the tip is usually in the marrow cavity. 3 At that point, the stylet should be removed and a 10- to 12-ml syringe (previously rinsed with 2% EDTA solution) attached to the needle. Vigorous negative pressure is applied by rapidly pulling on the plunger of the syringe. To avoid dilution with peripheral blood, aspiration should be stopped when the first drop of marrow appears in the syringe. Failure to obtain marrow may be due to improper placement of the needle, plugging of the needle, myelofibrosis, or marrow that is packed with neoplastic cells. Replacing the stylet and slightly repositioning the needle should be tried. If repositioning fails to obtain a marrow specimen, aspiration should be attempted at another site. PREPARATION OF BONE MARROW SPECIMENS Preparation of high-quality bone marrow aspiration smears is essential for cytologic evaluation. Bone marrow specimens clot rapidly after collection. To permit time for preparing several high-quality smears, the syringe should be rinsed with 2% EDTA solution. Once marrow is obtained, the syringe should be removed from the needle and a drop of marrow expressed onto a microscope slide (Figure 4). The drop should contain blood, fat droplets (seen as a fatty sheen to the surface of the drop), and variable numbers of small, white flecks (i.e., unit particles) that represent pieces of marrow tissue. To prepare smears for cytologic evaluation, a spreader slide is touched to the drop of marrow and then picked up, and a wedge-type smear is made on another clean slide (i.e., similar to preparing blood smears). In this way, 10 to 20 slides can be made from a single drop of marrow. The slides should be rapidly dried and then inspected. A properly made smear should be thin, contain fat droplets (round clear areas), and have several white specks (i.e., unit particles) that are frequently located on the feathered edge. If unit particles are not visible on the slides, any unit particles that are present in the marrow specimen should be picked out and placed on a clean slide and a second slide placed on top. The two slides should then be pulled apart vertically to create a squash preparation. The wedge-type smears are thin enough to provide good cell morphology and, in most cases, provide enough unit particles to assess cellularity. Squash preparations tend to be thick, contain many lysed cells, and may stain very darkly, making assessment of cell morphology difficult. The quality of the cytologic preparation can be assessed in several ways. The presence of fat or unit particles in the marrow sample indicates that the needle was in the marrow cavity. However, unit particles may not be present in poorly cellular or fibrotic marrow specimens. If fat or unit particles are present, poorly cellular samples usually indicate either low marrow cellularity or dilution of marrow with blood. Marrow smears should be stained and examined microscopically to determine whether slides are cellular and cells are intact. Once adequate marrow aspiration smears have been obtained, slides can be stained and examined within 24 hours or fixed in absolute methanol for 5 seconds (the

4 674 Small Animal/Exotics Compendium September 2002 Figure 5 Jamshidi needle placed in the iliac crest of a dog. Figure 6 Unit particles in an aspiration smear of dog bone marrow. Note the presence of fat (clear areas) and hemic cells (dark blue areas). (Wright s stain, original magnification, 40) fixative for most rapid Wright s stains is absolute methanol). Slides can be stained with new methylene blue or a rapid Wright s-type stain. Bone marrow needs to be stained for approximately twice as long as blood smears when Wright s-type stains are used. It is important not to expose unstained bone marrow aspiration smears to formalin or formalin fumes. Even brief exposure results in a marked decrease in staining quality when Wright s-type stains are applied. CORE BIOPSY TECHNIQUE A concurrent bone marrow core biopsy is recommended in addition to bone marrow aspiration. Core biopsy specimens are primarily used to assess cellularity and stromal alterations. 2 3 Without core biopsies, stromal alterations, including myelofibrosis, myelonecrosis, metastatic neoplasia, and inflammation, may be missed. Although all three sites described for obtaining aspirates have been used, core biopsies are most frequently obtained from the wing of the ilium. 2 With the dog in lateral recumbency, the Jamshidi needle is inserted lateral to the iliac crest (Figure 5). The needle path is from lateral to medial. The needle is initially advanced with the stylet in place until the needle begins to enter the bone. The stylet is removed, and the needle is advanced slowly with rotation until it exits the medial side of the ilium. The needle tip can be palpated if the dog is thin. The needle is then rotated several times to make sure all connections of the specimen are detached and is removed with continuous rotation. The biopsy specimen is pushed out of the needle from the cutting end using a probe. The biopsy specimen should be immediately placed into 10% neutral buffered formalin. When submitted to a clinical laboratory, biopsy tissue is embedded in paraffin, sectioned, and stained before examination. EVALUATION OF BONE MARROW ASPIRATION SMEARS Bone marrow aspiration smears cannot be adequately evaluated without knowledge of concurrent changes occurring in the blood. Therefore, blood should be collected concurrently with the bone marrow sample and a CBC or blood smear evaluation conducted. Evaluation of bone marrow aspiration specimens includes assessment of cellularity; the relative number and morphology of megakaryocytes present; the myeloid: erythroid (M:E) ratio; maturation within the erythroid and myeloid series; erythroid and granulocyte morphology; the number and morphology of lymphocytes, plasma cells, monocytes, and macrophages present; and the amount of iron (i.e., hemosiderin) present. 1,2 Cellularity Overall cellularity of the aspiration smear is relatively easy to judge as increased, decreased, or normal. However, hypocellular smears may be misleading due to blood contamination during aspiration. To more precisely evaluate cellularity, unit particles should be examined (Figure 6). The ratio of fat (seen as round, clear spaces) to hemic cells (seen as cell clusters or reddishblue stained masses) provides an estimate of cellularity. Many unit particles should be examined because cellularity may vary. In normal cellularity marrow, unit particles should contain 25% to 75% hemic cells. 2 However, this ratio may vary with the age of the animal and the site of aspiration. Bone marrow cellularity decreases throughout life. In young animals, unit particles should be 50% to 75% hemic tissue, whereas in older dogs, unit particles should be 25% to 50% hemic tissue. After judging the cell density in the smear and the cellularity of unit particles, overall marrow cellularity should be estimated as increased, decreased, or normal.

5 Compendium September 2002 Canine Bone Marrow Evaluation 675 Table 1. Terms Used To Describe Normal and Abnormal Morphology in Canine Bone Marrow Erythroid cells in marrow Immature Mature Granulocyte cells in marrow Immature Mature Production of hematopoietic cells Erythropoiesis Granulopoiesis Thrombopoiesis Marrow cellularity Hyperplasia Hypoplasia Aplasia Myelodysplasia Dyserythropoiesis Dysgranulopoiesis Dysmegakaryopoiesis Myelodysplastic syndrome Leukemia Rubriblast, prorubricyte Rubricyte, metarubricyte, reticulocyte Myeloblast, promyelocyte Myelocyte, metamyelocyte, band, segmenter Production of erythroid cells Production of neutrophils, eosinophils, and basophils Production of platelets (also called megakaryocytopoiesis) Increased cellularity Decreased cellularity Very few or no cells Dysplastic features in erythroid cells Dysplastic features in granulocytes Dysplastic features in megakaryocytes General term for clinical disorders characterized by dysplastic cells in bone marrow; three types have been described Malignancy of hemic cells (acute leukemias have >30% blast cells present) Megakaryocytes Megakaryocytes are easily identified using the 4- or 10-times objective. These cells are 10 to 20 times the diameter of other hemic cells, have multilobulated nuclei, and are usually concentrated on the feathered edge of the smear and around unit particles. Megakaryocyte numbers vary from slide to slide. In general, 50 to 100 megakaryocytes are present on wedge-type smears with a normal number of megakaryocytes. Because of the low number of megakaryocytes expected and variable distribution, at least three smears should be examined before megakaryocyte numbers are considered to be decreased. In normal marrow, greater than half the megakaryocytes are mature. 2 Mature megakaryocytes have multilobulated nuclei with 32 or 64 nuclei and a light blue or pink granular cytoplasm, whereas immature megakaryocytes have fewer nuclear lobulations and a dark blue, relatively homogeneous, cytoplasm. 2 Differential Cell Count Bone marrow contains a large number of cell types representing the developmental stages of erythroid and myeloid cells (Figure 7). The morphology of each developmental stage of the erythroid and myeloid series, as well as other cells in canine marrow, has been reviewed. 2,6 Conducting a differential cell count involves categorizing at least 20 different cell types. At least 500 cells should be counted. Because of the difficulty and time required to complete differential cell counts, cell distribution is usually estimated. However, a modified approach that speeds and simplifies the process of differential cell counting can be used. Instead of enumerating each developmental stage of erythroid and myeloid cells, cells are classified as immature or mature myeloid or immature or mature erythroid (Figure 7). Lymphocytes and monocyte/ macrophages are also quantified. Mature erythroid cells (i.e., rubricytes, metarubricytes) can be differentiated from immature erythroid cells (i.e., rubriblasts, prorubricytes) by the smaller size of the immature cells and their gray to pink cytoplasm, indicating the incorporation of hemoglobin. Mature granulocytes (i.e., metamyelocytes, bands, segmenters) can be differentiated from immature granulocytes (i.e., myeloblasts, promyelocytes) by the indented or lobed nucleus of mature granulocytes. With this approach, 500-cell differential cell counts can be accomplished in less than 5 minutes.

6 676 Small Animal/Exotics Compendium September 2002 Possible Cell Types in a Differential Cell Count Figure 7 Bone marrow aspiration smear from a healthy dog. Notice the presence of immature erythroid cells (red arrow) and immature myeloid cells (blue arrow). Immature erythroid cells can be differentiated from immature myeloid cells by their smaller size and darker-staining nucleus and cytoplasm. Also notice the presence of mature erythroid cells (black arrow) and mature myeloid cells. Mature erythroid cells are characterized by their smaller size, clumped nuclear chromatin, and gray to pink cytoplasm. Mature myeloid cells (yellow arrow) are characterized by indented or lobed nuclei. Also notice that the number of mature erythroid and myeloid cells exceeds the number of immature erythroid and myeloid cells (Wright s stain). Myeloid:Erythroid Ratio The M:E ratio is a comparison of the number of myeloid cells (i.e., granulocytes, monocytes) to the number of nucleated erythroid cells. Lymphocytes, plasma cells, macrophages, and nonnucleated erythrocytes are not included. The M:E ratio can be determined by completing a 500- or 1000-cell differential cell count or estimated by determining the relative number of myeloid and erythroid cells in 10 or more fields in various areas of the smear. 2 Microscopic fields containing unit particles should be avoided because of the difficulty in differentiating cells and the presence of a large percentage of myeloid cells in these areas. Myeloid cells can be differentiated from erythroid cells by cell size, nuclear color, and nuclear shape (Figure 7). Normal M:E ratios vary from 1:1 to 2:1; however, the appropriateness of any M:E ratio must be interpreted in consideration of the overall hematologic condition. 2 Rubriblasts Prorubricytes Rubricytes Metarubricytes Myeloblasts Promyelocytes Neutrophil myelocytes Eosinophil myelocytes Basophil myelocytes Neutrophil metamyelocytes Eosinophil metamyelocytes Basophil metamyelocytes Band neutrophils Segmented neutrophils Segmented eosinophils Segmented basophils Lymphocytes Plasma cells Monocytes Macrophages Maturation of Granulocytes and Erythroid Cells The maturation sequence of the granulocytic and erythroid series should be evaluated. In normal marrow, more than 90% of cells in the erythroid series (i.e., rubricytes, metarubricytes) and more than 80% of cells in the granulocytic (i.e., metamyelocytes, bands, segmenters) series are relatively mature (Figure 7). 7 As previously noted, mature erythroid cells can be differentiated from immature erythroid cells by the presence of a gray to pink cytoplasm associated with the incorporation of hemoglobin in immature cells. Mature granulocytes can be identified by their indented or lobed nuclei. With this approach, it is possible to determine the granulocyte maturation index (mature granulocytes/ immature granulocytes) and erythroid maturation index (mature erythroid cells/immature erythroid cells). 7 9 Normal granulocyte maturation index is 4.4 ± 1.4, and normal erythroid maturation index is 5.2 ± Atypical Cells Atypical cells include increased numbers of blast cells and cells with abnormal morphology. Blast cells are characterized as poorly differentiated, immature basophilic cells with dispersed nuclear chromatin that frequently have nucleoli and may have a Golgi zone. Normally, these cells are less than 1% of total nucleated marrow cells. Blast cells may increase in cases of myelodysplastic syndromes (5% to 30% of all nucleated cells) and acute leukemias (more than 30% of all nucleated cells). 10 Cells with atypical morphology are called dysplastic. The presence of dysplastic erythroid cells is called dyserythropoiesis, of dysplastic myeloid cells is called dysgranulopoiesis, and of dysplastic megakaryocytes is called dysmegakaryopoiesis. Miscellaneous Cells Miscellaneous cells include monocytes, macrophages, lymphocytes, plasma cells, osteoblasts, and osteoclasts. 1,2,6 Lymphocytes in bone marrow are usually small, similar to those seen in blood. An increased number of mature plasma cells in the bone marrow is usually an indication of a systemic immune response. Most frequently associated disease conditions include immune-mediated hemolytic anemia, immune-mediated thrombocytopenia, and rickettsial infections. 1 A large increase in plasma

7 Compendium September 2002 Canine Bone Marrow Evaluation 677 cells or the presence of large or atypical plasma cells is consistent with multiple myeloma. Although plasma cells typically make up less than 5% of total nucleated marrow cells, they can be up to 15% of the total in healthy dogs. Monocytes and macrophages usually make up less than 2% of total nucleated cells in marrow. Osteoclasts and osteoblasts are rarely found. Iron Iron is present in marrow as soluble aggregates (i.e., ferritin) in developing erythroid cells and as large insoluble masses (i.e., hemosiderin) in macrophages. 2 Hemosiderin appears as greenish-black, amorphous masses within macrophages or free granules in the background. Hemosiderin is most prominent in unit particles. The amount of hemosiderin present is subjectively graded as absent, present, or increased. To further quantify iron content, a Prussian blue stain, which stains iron a deep blue, can be applied. 2 SPECIAL TECHNIQUES Several special diagnostic techniques have been applied to bone marrow. Immunocytochemical techniques and flow cytometry have been used to identify the lineage of leukemia cells. Immunocytochemical stains detect cell lineage-restricted enzymes in granulocytes and monocyte/macrophages and to a lesser degree in lymphoid cells. 11,12 Flow cytometers are instruments in which cells in suspension are passed single file through a laser beam. The forward-angle light scatter properties are used to determine cell size, and the sideangle light scatter properties are used to determine cell granularity. 13,14 Results are displayed on a computer as side-scatter versus forward-scatter dot plots or contour SSC-H F1: R3 R1 R FSC-H Figure 8 Flow cytometric side-angle (SSC-H) versus forwardangle (FSC-H) light scatter plot of normal canine bone marrow. A template was used to identify mature (R1) and immature (R2) erythroid cells, band and segmented neutrophils (R3), immature granulocytes (R4), and metamyelocytes (R5). R5 R4 plots (Figure 8). Flow cytometry can also be used to identify specific cell types through the application of fluorescence-labeled monoclonal antibodies. 14 A battery of monoclonal antibodies can be used to determine the lineage of leukemic cells. CLASSIFICATION OF BONE MARROW DISORDERS Conclusions based on bone marrow evaluation can be descriptive of the morphologic changes or tailored to the specific case. Clinical pathologists frequently have only bone marrow aspiration smears and core biopsy specimens (and perhaps the age, sex, and breed of the dog) on which to base their conclusions. With this data, a morphologic conclusion that usually combines several of the terms listed in Table 1 can be stated. 1,2 For example, if the cellularity is increased (i.e., hyperplasia) and the M:E ratio is increased, an appropriate conclusion would be myeloid hyperplasia. Alternatively, if cellularity is decreased (i.e., hypoplasia) and the M:E ratio is increased, an appropriate conclusion would be erythroid hypoplasia. Even these general conclusions may be inaccurate if peripheral blood data have not been evaluated. A more definitive conclusion can frequently be made if the case history and CBC results are available. For example, if bone marrow cellularity and the M:E ratio are normal, the hematocrit is 14%, the reticulocyte count is 0%, and the case history indicates that the dog bled severely 24 hours before hematologic evaluation, the conclusion would be peracute blood loss with inadequate time for the bone marrow to respond. Alternatively, if bone marrow cellularity is increased (i.e., hyperplasia), the M:E ratio is increased, the hematocrit is 28%, the reticulocyte count is 0%, and the animal had a history of chronic pneumonia, a diagnosis of anemia secondary to inflammatory disease would be appropriate. These examples indicate that additional conclusions can be reached when bone marrow alterations are interpreted with knowledge of the case history and changes in peripheral blood cell counts. REFERENCES 1. Harvey JW: Atlas of Veterinary Hematology: Blood and Bone Marrow of Domestic Animals. Philadelphia, WB Saunders Co, 2001, pp Harvey JW: Canine bone marrow: Normal hematopoiesis, biopsy techniques, and cell identification and evaluation. Compend Contin Educ Pract Vet 6: , Relford RL: The steps in performing bone marrow aspiration and core biopsy. Vet Med 86: , Weiss DJ, Klausner JS: Drug-induced aplastic anemia in dogs: Eight cases ( ). JAVMA 196: , Deldar A, Lewis H, Bloom J, Weiss L: Cephalosporin-induced changes in the ultrastructure of canine bone marrow. Vet Pathol 25: , Reagan WJ, Sanders TG, DeNicola DB: Veterinary Hematology Atlas

8 678 Small Animal/Exotics Compendium September 2002 of Common Domestic Species. Ames, Iowa State University Press, 1998, pp Guelfi JF, Trumel C: Bone marrow maturation indexes: A retrospective study in 256 dogs and 61 cats. Vet Clin Pathol 30: , Weiss DJ, Lulich J: Myelodysplastic syndrome with sideroblastic differentiation in a dog. Vet Clin Pathol 28:59 63, Canfield PJ, Watson ADJ: Investigations of bone marrow dyscrasia in a poodle with macrocytosis. J Comp Pathol 101: , Raskin RE: Myelopoiesis and myeloproliferative disorders. Vet Clin North Am Small Anim Pract 26: , Grindem CB, Stevens JB, Perman V: Cytochemical reactions in cells from leukemic dogs. Vet Pathol 23: , Facklam NR, Kociba GJ: Cytochemical characteristics of leukemic cells from 20 dogs. Vet Pathol 22: , Weiss DJ: Use of monoclonal antibodies to refine flow cytometric differential cell counting of canine bone marrow cells. Am J Vet Res 62: , Weiss DJ: Evaluation of proliferative disorders in canine bone marrow by use of flow cytometric scatter plots and monoclonal antibodies. Vet Pathol 38: , Ta ARTICLE #1 CE TEST ble The 1. Terms article Used you have to Describe read qualifies Normal for and 1.5 Abnormal contact hours of Continuing Education Credit from Morphology in Canine Bone Marrow sible Cell Types in a Differential the Auburn University College of Veterinary Medicine. Choose the best answer to each of the following questions; then mark your answers on the postage-paid envelope inserted in Compendium. C 1. Which statement regarding bone marrow aspiration technique is incorrect? a. General anesthesia is usually needed. b. The site should be aseptically prepared. c. Illinois sternal needles are commonly used. d. The syringe should be rinsed with EDTA solution. e. Sterile gloves should be worn. 2. Which site for bone marrow aspiration is most accessible in obese dogs? a. iliac crest d. rib b. proximal femur e. ischial tuberosity c. proximal humerus 3. Which condition is not a good indication for performing a bone marrow biopsy? a. nonregenerative anemia b. persistent neutropenia c. unexplained hypercalcemia d. atypical cells in the blood e. regenerative anemia with large numbers of spherocytes in the blood 4. Which statement about bone marrow aspirates is incorrect? a. The presence of lipid indicates that the needle was in the marrow cavity. b. At least 1 ml of marrow should be aspirated. c. The presence of unit particles indicates that the needle was in the marrow cavity. d. The syringe can be rinsed with 2% EDTA solution to prevent clotting. e. Marrow aspiration smears should not be exposed to formalin or formalin fumes. 5. Which characteristics of immature erythroid cells differentiate them from immature myeloid cells? a. smaller size and basophilic cytoplasm b. larger size and basophilic cytoplasm c. smaller size and pink cytoplasm d. smaller size and lobed nuclei e. larger size and granular cytoplasm 6. Which characteristics of mature erythroid cells differentiate them from mature myeloid cells? a. smaller size and dispersed chromatin pattern b. larger size and pink cytoplasm c. larger size and clumped chromatin pattern d. smaller size and clumped chromatin pattern e. larger size and granular cytoplasm 7. Bone marrow characterized by increased cellularity, normal numbers of megakaryocytes, increased M:E ratio, normal morphology and maturation in the erythroid and myeloid series, and neutrophilia and left shift in the peripheral blood is most consistent with which condition? a. myeloid hyperplasia b. erythroid hypoplasia c. leukemia d. myeloid hypoplasia e. erythroid hyperplasia 8. Bone marrow characterized by increased cellularity, normal number of megakaryocytes, decreased M:E ratio, normal morphology and maturation in the erythroid and myeloid series, and regenerative anemia in the peripheral blood is most consistent with which condition? a. myeloid hyperplasia b. erythroid hypoplasia c. leukemia d. myeloid hypoplasia e. erythroid hyperplasia 9. Bone marrow characterized by decreased cellularity, normal numbers of megakaryocytes, decreased M:E ratio,

9 Compendium September 2002 Canine Bone Marrow Evaluation 679 normal morphology and maturation in the erythroid and myeloid series, and neutropenia with no left shift in the blood is most consistent with which condition? a. myeloid hyperplasia d. myeloid hypoplasia b. erythroid hypoplasia e. erythroid hyperplasia c. leukemia 10. Bone marrow characterized by increased cellularity, normal number of megakaryocytes, increased M:E ratio, normal morphology and maturation of erythroid cells, atypical myeloid cells, 10% blast cells, and neutropenia in the peripheral blood is most consistent with which condition? a. myeloid hypoplasia b. erythroid hypoplasia c. leukemia d. dysgranulopoiesis e. dyserythropoiesis