1. Evacuated Tubes and Additives for Blood Specimen Collection (NCCLS, 1999 ) 2000 년 5 월 4 일 이 운 형

2. Materials 1Tube (tube shall) made from material that allows an adequate, undistorted view of the contents sufficient physical strength to withstand normal use capable of retaining vacuum for the stated shelf life available in glass and plastic.

3. Materials 2Construction All materials used in the construction of the container and any lubricants, coatings, or additives must not adversely influence the results obtained when the container is used as intended. 3Closure Material provide for needle puncture and reseal hold vacuum for the stated shelf life permit easy removal of the closure

4. Materials 4Specifications chemical additives must either meet the specifications of those compendia or package labeling shall state where they differ.

5. Figure 1. Standard Double-ended Blood-collection Needle/holder.

6. Sizes 1Tube Dimensions The nominal tube size the external tube diameter and overall length, the maximum quantity of specimen to be drawn into the tube

7. Typical Sizes Are As Follows: Nominal Size(mm)Maximum Draw (mL) Glass Maximum Draw(mL) Plastic 10 x 653- 13 x 7554 13 x 10076 16 x 75710 16 x 10010- 16 x 1251514

8. Sizes 2Compatibility with Centrifuge Carriers The complete container shall fit commonly used centrifuge carriers without breakage during proper placement, spinning, or removal from the centrifuge

9. Design 1Complete Container suitable for use on mechanical mixers, including roller-type mixers The closure : not become loose during mixing when tested in accordance with specified methods

10. Design 2Closure It can be easily removed by gripping with gloved fingers or using mechanical extractors. It is not necessary during removal to touch any inside tube surface or any closure surface that is exposed to the contents of the tube. It will not be removed or loosened when used to collect a blood specimen with a blood-collection needle and holder.

11. Construction 1Strength A complete container (with the specimen) must be capable of withstanding the centrifugal force resulting from an acceleration of 2,200 x g relative centrifugal force (RCF)

12. Construction 2Exterior Texture No part of the container shall have a sharp edge, projection, or surface roughness capable of cutting, puncturing, or breaking the skin of the user

13. Additives Table 1 The additives with their associated "initial" identification and color coding and the suitable salts and their physical states. The additives were chosen for their ready ability to mix with the blood specimen, and tolerances are given for the quantity to be used per milliliter of whole blood.

14. Table 1

20. Appendix A - EDTA Desirable for use in the hematology laboratory because cellular components of the blood are preserved

21. Appendix A - EDTA Chelation activity takes place : With calcium and other divalent ions : With calcium and other divalent ions, which can act as enzyme cofactors  Not suitable for calcium, iron, alkaline phosphatase, creatinine kinase, and leucine aminopeptidase determinations  Not suitable for calcium, iron, alkaline phosphatase, creatinine kinase, and leucine aminopeptidase determinations.

22. A2Description - EDTA One of a class of aminopolycarboxylic acids

23. A2.1Mode of Action - EDTA Chelate effectively calcium in blood Calcium : necessary in the coagulation cascade,  Inhibits and stops the series of intrinsic and extrinsic events, which cause clotting.  Inhibits the conversion of prothrombin to thrombin, and the action of thrombin on fibrinogen to form fibrin

24. A2.2Anticoagulant Properties - EDTA Unacceptable anticoagulant for coagulation tests, except for the isolation and examination of platelets. Note that, in some patients, platelets can aggregate in the presence of EDTA because of the occurrence of platelet- specific antibodies.

25. Appendix A – EDTA MWFormSolubilitychelation value EDTA, Free Acid292.2White powder- odorless Saturated solution (20 °C), 200 to 300 mg/L. 1 g or 3.4 mmol EDTA (acid) chelates 3.4 mmol or 136 mg of calcium ion. EDTA Na2 · 2H2O372.2White crystalline powder-odorless Approximately 100 g/L at 20 °C. 1 g or 2.6 mmol EDTA (disodium salt) chelates 2.6 mmol or 105 mg of calcium ion. EDTA K2 · 2H2O404.4White crystalline powder-odorless Approximately 1650 g/L at 22 °C. 1 g or 2.4 mmol EDTA (dipotassium salt) chelates 2.4 mmol or 100 mg of calcium ion. EDTA K3406Clear, odorless liquid approximately 1650 g/L at 22 °C 1 g or 2.4 mmol EDTA (tripotassium salt) chelates 2.4 mmol or 100 mg of calcium ion

26. A3Required Quantity - EDTA A3.1coagulation prevention Enough EDTA must be present to prevent coagulation But excessive amounts cause morphological changes in blood cells.

27. A3Required Quantity - EDTA A3.2EDTA salts The amount of EDTA salt added to blood should be 4.55 ± 0.85 mmol/ml of blood.

28. A3Required Quantity – EDTA A3.2EDTA salts EDTA Saltmg/mL Blood Disodium EDTA dihydrate EDTA Na2 · 2H2O 1.4 – 2.0 Dipotassium EDTA dihydrate EDTA K2 · 2 H2O 1.5 – 2.2 Tripotassium EDTA anhydrous EDTA K3 1.5 – 2.2

29. A3Required Quantity – EDTA A3.2EDTA salts The tripotassium salt is usually used in liquid form. The dilution effect with blood can give different results on tests when there is less than a complete blood draw. Packed cell volume determinations can also be lower with the tripotassium salt for the same reason.

30. A4Selection of EDTA EDTA is available as free acid, as well as the disodium, dipotassium, and tripotassium salts. The salts are preferable for use due to a high solubility.

31. A4Selection of EDTA Regardless of the form or type of EDTA salt used, all tubes should be inverted several times (8 to 10 times) to ensure thorough mixing and, therefore, proper anticoagulation.

32. A4Selection of EDTA Disodium and dipotassium salts are commonly used in the dry form Tripotassium EDTA is normally used in the liquid form to further enhance the anticoagulant activity. The tripotassium EDTA solutions commonly used cause a 1 to 2% dilution of the blood.

33. A4Selection of EDTA Dipotassium EDTA is used with certain devices. : anticoagulant of choice in specimen collection for blood collection for blood cell counting and sizing.

34. A5Stability of the Specimens – EDTA Users should determine the stability of the specimens. Stability of specimens can vary depending on the reagents, system, instrumentation, and storage conditions.

35. A5Stability of the Specimens – EDTA "short draw" : a principal hazard to stability It is strongly recommended that the proper amount of blood be drawn into tubes for appropriate and safe anticoagulation. Having a great excess of EDTA in the blood can shrink red cells because the hypertonicity of the plasma with increased ionic concentration.

36. A5Stability of the Specimens – EDTA A ratio of 1.5 mg EDTA/ml of blood is considered optimal Blood and EDTA must be in balance. A ratio of 1.5 mg EDTA/ml of blood is considered optimal; Larger quantities can affect red cell size.

37. A6Labeling Claims – EDTA EDTA-containing devices must be labeled properly to provide users with sufficient data for correct use. It is recommended that manufacturers provide the following data: Salt of EDTA Weight and/or volume of EDTA Volume of blood required Expiration date Storage conditions.

38. Appendix B - Heparin Minimal chelating properties Minimal effects on water shifts Relatively low cation concentration

39. Appendix B - Heparin Sodium and ammonium salts of heparin are available. Lithium heparin is recommended as an anticoagulant Because it is least likely to interfere when performing tests for other ions.

40. Appendix B - Heparin Should be used in a device for the determination of ph, blood gases, electrolytes, and ionized calcium Should not be used when coagulation tests are to be performed.

41. B1Definition - Heparin B1.1description A sulfated glycosaminoglycan that prolongs the clotting time of blood. Obtained from bovine and porcine lungs and intestines.

42. B1Definition - Heparin B1.1description The potency of lithium heparin, calculated on a dried basis, should be not less than 140 USP heparin units/mg It should be within 10% of the potency stated on the label. Lithium heparin is essentially free of extraneous ions. It should not be used for collection of blood for lithium levels.

43. B1Definition - Heparin B1.2 mode of action Acts primarily through a complex that it forms with antithrombin III. This complex accelerates the inhibition of thrombin and activated factor X to prevent clotting or activation of thrombin.

44. B3Quantity Required - Heparin Manufactured devices usually contain an excess of heparin. A recommended range : 10 to 30 USP units/ml of blood

45. B4Labeling Claims - Heparin It is recommended that manufacturers provide the following data: Salt of heparin USP strength of heparin (anhydrous weight) Volume of blood required Expiration date Storage conditions Methods of blood collection and mixing.

46. Appendix C - Sodium Citrate The anticoagulant of choice for coagulation tests.

47. C1Description-sodium Citrate : A tricarboxylic acid. : A molecular weight of 192. The trisodium salt The usual form of use Molecular weight of 294 The sodium acid citrate (disodium citrate, disodium hydrogen citrate) is also used “ Buffered" sodium citrate The combination of sodium citrate and citric acid

48. C2Mode of Action-sodium Citrate Primarily to chelate, or bind, calcium The binding effect of citrate can be reversed by recalcifying the blood or derived plasma to its normal state.

49. C2Mode of Action-sodium Citrate This simple reversible action makes it highly desirable for coagulation determinations. The coagulation factors are not affected by citrate and its effect on cells and platelets is minimal.

50. C5Use - Sodium Citrate Citrate tubes containing sodium citrate or buffered sodium :Contain a 3.8% solution Citrate tubes containing of the dehydrated form of trisodium citrate :Contain a 3.2% solution

51. C5Use - Sodium Citrate 1Anticoagulant The anticoagulant used for coagulation assays should be 105-109 mmol/L, 3.13%- 3.2% (commonly described as 3.2%) of the dihydrate form of trisodium citrate (Na3C6H5O7 x 2H 2O), buffered or nonbuffered.

52. C5Use - Sodium Citrate 2Anticoagulant/Blood Ratio Blood : the sodium citrate dihydrate = 9:1 Inadequate filling of the collection device will decrease this ratio, and may lead to inaccurate results.

53. C5Use - Sodium Citrate 3Citrate Concentration Adjustments The final citrate concentration in the blood should be adjusted in patients who have hematocrit values above 0.55 (55%).

54. C5Use - Sodium Citrate

55. The End