Venipuncture Procedures: Sources of Human Peripheral Blood Cells, Serum, and Plasma

Barbara E. Bierer1, Warren Strober2

1 Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts, 2 National Institute of Allergy and Infectious Diseases, Bethesda, Maryland
Publication Name:  Current Protocols in Immunology
Unit Number:  Appendix A.3F
DOI:  10.1002/cpim.27
Online Posting Date:  August, 2017
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Peripheral blood is the chief source of mononuclear cells (lymphocytes and monocytes) for immunologic studies of humans. This appendix presents protocols for obtaining blood by simple venipuncture when relatively small amounts of blood (10 to 100 ml) are necessary or by lymphapheresis when large amounts (300 to 5000 ml) are necessary. Cells collected by these procedures can be further separated by techniques described in Chapter 7. © 2017 by John Wiley & Sons, Inc.

Keywords: lymphapheresis; peripheral blood; plasma; serum; venipuncture

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Table of Contents

  • Introduction
  • Basic Protocol 1: Blood Collection by Venipuncture
  • Alternate Protocol 1: Blood Collection by Lymphapheresis
  • Literature Cited
  • Tables
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Basic Protocol 1: Blood Collection by Venipuncture

  • IRB/REC approved clinical research protocol for research procedures
  • Human volunteer donor who has read, understood, and signed an informed consent document, unless waiver of documentation of consent is approved by the IRB/REC
  • Heparin, preservative‐free (optional)
  • 70% alcohol wipes or dry sterile gauze pads, dry and soaked in 70% isopropanol
  • Multiple approaches for obtaining blood can be utilized, including:
  • Vacutainer needles (typically 20‐ or 21‐G) and Vacutainer needle holders or equivalent (e.g., Scientific Products), and Vacutainers, type depending on the purpose of collection (Table 3.0.1)
  • Appropriately sized syringe (10‐, 30‐, 50‐ml) with 17‐ to 21‐G and 23‐G “safety” needles
  • Appropriately sized syringe (10‐, 30‐, 50‐ml) or Vacutainer with “butterfly” needles
  • For collection of blood using non‐Vacutainer needle and syringe, plastic or glass blood collection tubes (e.g., Scientific Products), type depending on the purpose of collection (see Table 3.0.1)
  • Well fitting, non‐sterile, latex or non‐latex gloves
  • Tourniquet
  • Sterile gauze pad
  • Sterile adhesive bandage
  • Puncture‐resistant, leak‐proof sharps container (see appendix 3P)
  • Zip‐lock bags for transport
  • Styrofoam containers, as required
  • Labels, as appropriate
Table 0.f.1   MaterialsRecommended Order of Draw, Color Coding, and Uses of Vacuum Blood Tubes (Vacutainers) and Non‐Vacutainer Blood Collection Tubes

Blood draw order Typical color of tube top a Additive Uses
1 Not applicable Broth Blood culture bottle; microbiology
2 Light blue Sodium citrate (reversible anticoagulant) Coagulation tests (e.g., PT, PTT), removes calcium from blood
3 Red top Additive‐free; clot forming Blood clots, serum for chemistries, immunology, blood bank
4 Red‐gray or gold Gel at bottom with clot activator Serum for chemistries, immunology, blood bank
5 Dark green Sodium or lithium heparin Inactivates thrombin and thromboplastin; used for plasma determinations and mononuclear cells after Ficoll‐Hypaque
6 Light green Lithium heparin and gel at bottom Anticoagulants (lithium‐based) separates plasma for plasma and chemistries
7 Purple EDTA (potassium salt) Whole blood. Removes calcium, for hematology, smears, blood bank
8 Pale yellow Acid‐citrate‐dextrose or sodium polyanethol sulfonate Complement inactivation, HLA tissue typing, DNA studies, blood cultures
9 Light gray Sodium fluoride and potassium oxalate Antiglycolytic, glucose measurements

 aVerify color code with manufacturer prior to use. Similar color coding is used for Vacutainer tubes and for sample collection tubes used for collection of blood that has been drawn with a needle and syringe.
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Literature Cited

  Fuss, I. J., Kanof, M. E., Smith, P. D., & Zola, H. 2009. Isolation of whole mononuclear cells from peripheral blood and cord blood. Current Protocols in Immunology, 85, 7.1.1–7.1.8. doi: 10.1002/0471142735.im0701s85.
Key References
  Estey, C. A., & Felder, R. A. (1997). Clinical evaluation of serial blood processing at point of care. Clinical Chemistry, 43, 360–362.
  Heller, M., & Krebs, C. (1997). Clinical handbook for health care professionals. Clifton Park, NY: Delmar Learning.
  NCCLS. (2007). Procedure for the collection of diagnostic blood specimens by venipuncture: Approved standard (6th ed.), (Vol. 27, No. 26. H3–A6). Wayne, PA: NCCLS.
Internet Resources
  BD Vacutainer Systems—preanalytical solutions. Becton, Dickinson, and Company Web site. 2004. Accessed 12 March 2017.
  OSHA Safety and Health Bulletin SHIB 03‐10‐15: Disposal of Contaminated Needles and Blood Tube Holders Used for Phlebotomy. Accessed 26 March 2017.
  Pathlab/LSR Specimen Collection Guidelines. 2015 version. Accessed 26 March 2017.‐eia_bloodlct.pdf
  RBP‐E1A: Collecting, Processing, and Handling Venous, Capillary, and Blood Spot Samples. 2005. Accessed 20 March 2017.‐guidelines‐on‐drawing‐blood‐best‐practices‐in‐phlebotomy‐Eng.pdf?ua=1
  WHO guidelines on drawing blood: Best practices in phlebotomy. Geneva: World Health Organization; 2010. Aaccessed 20 March 2017.
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