1. Hematology Unit 2 Chapter 7 Sample Collection and Handling Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved.

2. Sample Collection Determines specific test procedures Equipment Supplies Vessels Jugular most appropriate in most cases Timing Fasted Before a procedure Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 2

3. Collection Equipment Needle and syringe Needle – largest comfortable for animal Syringe must be closest to the sample size needed 2.5 mL needed, use 3 mL syringe Vacutainer Preferred method Multiple samples collected with one stick Composed of needle, needle holder, and collection tubes Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 3

4. Collection Equipment (cont.) Vacutainer tube selection Size Range from 1 microliter to 15 mL Choose correct size to minimize vein collapse Fill to correct volume Ensure anticoagulant is in proper ratio to blood Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 4

5. Whole Blood Use proper anticoagulant Once collected Gentle rocking to mix Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 5

6. Plasma and Serum Plasma Fluid portion of whole blood 90% water, 10% dissolved constituents Proteins, carbohydrates, vitamins, hormones, enzymes, lipids, salts, waste, antibodies, other ions, and molecules Serum Plasma without fibrinogen During clotting fibrinogen is converted to insoluble fibrin clot matrix The clot squeezes out the serum Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 6

7. Anticoagulants Required when whole blood or plasma is needed Choose appropriate anticoagulant for specific test If unable to run test within 1 hour – refrigerate; do not freeze Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 7

8. Anticoagulants (cont.) Heparin Use ratio of 20 U/mL of blood to be collected Small amounts Coat the inside of syringe Vacuum tubes are premeasured to proper amount Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 8

9. Anticoagulants (cont.) Ethylenediaminetetraacetic acid (EDTA) Preferred for hematologic studies Does not alter cell morphology Do not use in plasma samples for chemical assays 1 to 2 mg/mL of blood Excessive amounts cause cells to shrink and invalidates cell counts in automatic analyzers Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 9

10. Anticoagulants (cont.) Oxalates Sodium, potassium, ammonium, or lithium salts Potassium most commonly used May inhibit lactate dehydrogenase and alkaline phosphate activity Not used in assays for potassium Citrates Sodium or lithium salts Sodium most commonly used Interferes with sodium assays Prevent clotting by binding with the calcium Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 10

11. Anticoagulants (cont.) Sodium fluoride Glucose preservative 6 to 10 mg/mL of blood May be added to other samples as a glucose preservative 2.5 mg/mL of blood Interferes with many enzymatic tests Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 11

12. Sample Volume Dependent upon quantity of serum or plasma needed Well-hydrated animal (PCV 50%) 50% cells, 50% fluid 10 mL blood = 5 mL fluid Dehydrated animals (PVC 70%) 70% cells, 30% fluid 10 mL blood = 3 mL fluid Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 12

13. Collection Procedure Gather equipment and supplies Preparation site – shave if possible and clean site Occlude vein – restrainer or tourniquet Perform venipuncture with minimal tissue injury Prevents contamination and hemolysis Stabilize vein Insert needle bevel up – 30 degrees After collection – remove needle and apply pressure If Vacutainer – fill to capacity Restraint – valuable tool!! Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 13

14. Collection Procedure (cont.) If transferring from syringe to tube: Remove needle from syringe before transferring Gently invert any tubes containing anticoagulant to ensure proper mixing with blood Label Date and time Patient name and ID number Owner name Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 14

15. Multiple Samples Order of draw Vacutainers should be used and in this order Citrate first Plain red top Tiger top – serum separator Green – heparin Lavender – EDTA Gray – potassium oxalate or sodium fluoride Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 15

16. Summary Whole blood Plasma Serum Supplies Anticoagulants Collection procedure Order of draw Copyright © 2015 by Mosby, an imprint of Elsevier Inc. All rights reserved. 16