Chapter 12 Additional Hematologic Tests Hematology Unit 2 Chapter 12 Additional Hematologic Tests Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Reticulocytes Immature erythrocytes Polychromatic staining Supravital stain – New methylene blue – fresh Not Romanowsky stains Differing staining methods Cats – 2 forms of retics Aggregate – large clumps of reticulum Punctate – unique to cats – 2 to 8 small singular basophilic granules Normal 0.4% aggregate and 1.5% to 10% punctate Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Reticulocyte Count Expression of the percentage of RBCs that are reticulocytes Percentage per 1000 RBCs Oil-immersion lens Done on all anemic animals except horses Horses do not release retics Useful in determining bone marrow response Reticulocyte counts should be interpreted according to the degree of anemia, because fewer mature erythrocytes are present in anemic animals, and reticulocytes are released earlier and persist longer than they do in normal animals. Higher percentages may be seen in hemolytic anemia as compared with hemorrhagic types of anemia. Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Corrected Reticulocyte Count Multiply observed reticulocytes by the observed PCV and then divide by the normal PCV. Normal – dog 45%; Cat 35% Example A dog has a PCV of 15% and observed retic count of 15% 15 × 15/45 ‒ 5% Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Reticulocyte Production Index Another option Involves the maturation time values Divide the corrected retic percentage by the maturation time for the observed patient’s PVC (Reticulocyte maturation index) (Table 12-1, page 78) Example: corrected retic count is 5% and reticulocyte maturation index is 2.5 5/2.5 = 2 which indicates the patient is producing reticulocytes at 2 times the normal rate. Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Bone Marrow Evaluation Valuable tool for diagnosis and prognosis in specific cases The need for this test is determined by the findings of the differential blood count. Persistent unexplained pancytopenia, neutropenia, or thrombocytopenia and nonregenerative anemia Abnormal morphologic changes or unexplained immature cells Stages neoplastic diseases Diagnosis parasitic infections – Ehrlichiosis, leishmaniasis Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Bone Marrow Evaluation (cont.) Collection Aspiration or removal of bone marrow core Restraint, sedation, or local or general anesthesia Aseptic technique Supplies EDTA prepared syringes Bone marrow needle No. 11 scalpel blade About 12 slides Sutures (if needed) Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Bone Marrow Collection Aspiration biopsy Several sites – head of femur or humerus Aseptic Rotated and pressure applied – attach syringe and aspirate Core biopsy Sometimes better sample quality Use different sites if done in conjunction with aspiration biopsy Similar procedure no aspiration – rotate needle to cut piece of bone from the cortex Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Preparing Marrow Smears Multiple slides for various preparation techniques Must be made immediately if not mixed with EDTA (1 hour with EDTA) Aspirate sample prepared like blood smears Line smears, starfish smears, compression smears Rapidly air-dried Stain with Romanowsky-type stain Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Evaluation of Bone Marrow Films Systematic approach Must have differential WBC count from concurrent peripheral blood Normal Adult: 50% nucleated cells and 50% fat Juvenile: 75% nucleated cells to 25% fat Geriatric: 25% nucleated cells to 75% fat Acellular – Aplasia Hypercellular – Hyperplasia Hypocellular – Hypoplasia Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Cells in Bone Marrow Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Results M:E ratio Maturation index (left shift index) When present Stainable iron Increased mitotic figures Increased osteoblasts, osteoclasts, and mast cells Phagocytized material Metastatic cells Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Erythrocyte Sedimentation Rate The rate RBCs fall in their own plasma will be altered in some disease states Some automated systems but not common Manual method Calibrated tube filled with fresh EDTA-treated blood to the upper calibrated mark Place in rack – perpendicular to table Allow to sit undisturbed at room temperature After 60 minutes (20 in horses), record the level of the top of the RBC column Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Erythrocyte Osmotic Fragility Test of the ability to withstand hemolysis in varying concentrations of saline Not routinely performed unless in referral centers or research settings Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

Summary Reticulocyte counts Bone marrow sampling and preparation Reporting bone marrow results Erythrocyte sedimentation rate Erythrocyte osmotic fragility Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.