Table of contents
Acronyms and abbreviations 5
Introduction 6
Part 1: Processing and Handling Blood Specimens — Pre-analytical Phase 9
SOP 1: Processing and handling blood specimens — pre-analytical phase 11
Part 2: Sample and Result Transport 19
SOP 2: Preparing samples for transport 20
SOP 3: Transporting and receiving samples 25
Part 3: Sample and Result Management in the Post-Analytical Phase 27
SOP 4: Sample and result management in the post-analytical phase 28
SOP 5: Returning results following off-site sample analysis 31
Part 4: Internal Audit for Mid or Low Level Laboratories 35
SOP 6: Internal audit for mid or low level laboratories 36
Part 5: Writing a Laboratory Quality Manual 40
SOP 7: Writing a laboratory quality manual 42
Appendices 47
Appendix A: Dispatch Register template 48
Appendix B: Sample/Result Transport Manifest 49
Appendix C: Sample/Result Tracking Register template 50
Appendix D: Internal audit checklist for low level laboratories 51
Appendix E: Explanation of internal audit checklist for mid or low level laboratories 57
Appendix F: Audit score tracker 64
Appendix G: Internal audit report action plan 65
References and resources 67
Acronyms and abbreviations
BD / Becton DickinsonCD4 / Cluster of differentiation 4
CDC / U.S. Centers for Disease Control and Prevention
DOB / Date of birth
EDTA / Ethylenediamine tetra-acetic acid
EGPAF / Elizabeth Glaser Pediatric AIDS Foundation
EQA / External quality assurance
HBV / Hepatitis B virus
HIV / Human immunodeficiency virus
NA / Not applicable
QA / Quality assurance
Rpm / Revolutions per minute
SOP / Standard operating procedure
WHO / World Health Organization
Introduction
Who should use the templates for SOPs?
These SOPs are intended as a resource to support laboratory staff working in clinical laboratories conducting laboratory tests related to HIV testing, care, and treatment. These SOPs are intended to be used along with materials found on CDC’s International Laboratory-related Resource and Activity Directory webpage (http://wwwn.cdc.gov/dls/ila/default.aspx) to support country laboratory teams to develop appropriate local materials.
What is the purpose of these SOP templates?
SOPs provide a comprehensive framework that will assist laboratories in designing and implementing processes and procedures that ensure the provision of high quality laboratory services. To provide high quality laboratory services, laboratories must produce results that are accurate, reliable and timely. Additionally, laboratories must implement quality management systems to reduce laboratory errors throughout all stages of laboratory work (Figure 1).
Figure 1: Quality management
Quality management applies to all laboratory processesThese SOPs were developed to be adapted and utilized at national, regional and local laboratories. As new recommendations evolve, SOPs can help to develop, standardize and implement realistic local approaches to the delivery of new or expanded activities. SOPs will:
n Reduce laboratory errors that may impact patient care.
n Ensure procedures are conducted consistently by everyone in the laboratory.
n Ensure all laboratory procedures are conducted safely.
n Facilitate comprehension of technical information related to laboratory procedures and analyses.
n Provide laboratory staff step-by-step instructions for organizing and carrying-out laboratory procedures.
n Assist clinical laboratories to implement and comply with national guidelines and standards.
n Improve communication and working relationships between laboratories and clinics.
n Prepare new staff to work in the laboratory and reinforce standards, procedures and job expectations for existing staff.
n Provide a mechanism to evaluate laboratory operations, serve as checklist for supervisors to monitor laboratory operations, and provide a framework for supportive supervision.
n Provide guidance to laboratory quality improvement initiatives and quality management systems.
SOPs are “working documents” that should be revised and updated regularly to assure compliance with new standards and procedures.
Overview of templates in this SOP document
n Pre-analytical and post-analytical laboratory SOPS: While errors can occur in any stage of laboratory work (pre-analytical, analytical and post-analytical), the majority of errors in clinical laboratories occur in the pre-analytical stage, followed by the post-analytical stage.[i],[ii],[iii] The uneven distribution of laboratory errors suggests that improvement is needed in laboratory processes that occur before and after sample analysis. The following SOPs address select pre and post analytical processes that are common sources of error:
- Sample handling and processing
- Sample and result transport
- Result and post-test specimen management
n Quality management SOPs: A quality manual is a document that provides a comprehensive description of systems and processes that promote quality in all laboratory operations. Internal audits can be used to assess these quality systems and processes, and to identify areas for quality improvement. Quality systems must be integrated into all laboratory stages and operations to ensure the provision of high quality laboratory services. The SOPs in this document provide guidance for developing quality management tools that are applicable to all stages of laboratory work.
- Writing a laboratory quality manual
- Conducting an internal audit for mid or low level laboratories
How were the templates developed and reviewed?
“Standard Operating Procedures: Template for Laboratory Activities” was prepared by EGPAF technical staff in the US following extensive review of many excellent global documents. The final version has the advantage of additional review for completeness and technical accuracy by staff in the US and country offices throughout Africa. A complete list of the references reviewed to inform the development of these SOPs is provided.
How should the templates be used?
This SOP template was developed to be adapted and utilized at national and local laboratories to reduce time spent locally developing SOPs. As new laboratory testing procedures and recommendations evolve, SOPs can help to develop, standardize and implement realistic local approaches to the implementation of new or expanded services. Their use can facilitate seamless, uninterrupted, high quality laboratory testing.
Part 1: Processing and Handling Blood Specimens — Pre-analytical Phase
Definitionsn CD4: A large glycoprotein molecule found on the surface of T lymphocytes that serves as the receptor for HIV and indicates immunologic status.
n Chemistry: The determination of the chemical constituents of blood by assay in a clinical laboratory as part of a diagnostic protocol.
n EDTA vacutainer: Tubes spray-coated with EDTA (ethylenediamine tetra-acetic acid) and used for whole blood hematology determinations and immunohematology testing.
n Freeze-thaw cycle: Cycle of freezing samples and thawing samples. Frequent freezing and thawing may contribute to sample degradation.
n Hematology: A medical science that studies blood and blood forming organs.
n Hemolysis: The rupture or destruction of red blood cells, causing the release of hemoglobin.
n Heparin vacutainer: Tubes spray-coated with either lithium heparin or sodium heparin used for plasma determinations in chemistry.
n Glucose vacutainer: Tubes containing a glycolytic inhibitor used for glucose determinations on plasma.
n Plasma: The clear, yellowish fluid portion of blood, lymph, or intramuscular fluid in which cells are suspended. It differs from serum in that it contains fibrin and other soluble clotting elements.
n Serology: A medical science that deals with the detection of antigens orantibodiesin blood serum
n Serum: The clear yellowish fluid obtained upon separating whole blood into its solid and liquid components after it has been allowed to clot.
n Universal Precautions: A set of precautions designed to prevent transmission of HIV, hepatitis B virus (HBV), and other blood borne pathogens when providing first aid or health care. Under Universal Precautions, blood and certain body fluids of all patients are considered potentially infectious for HIV, HBV and other blood borne pathogens.
n Vacutainer: Glass tube with a rubber stopper in which air can be removed to create a vacuum usually used to draw blood. Vacutainer™ is a registered trademark of Becton, Dickinson and Company.
Safety and Precautions
n All individuals handling samples should receive biosafety training.
n Follow Universal Precautions (http://www.cdc.gov/niosh/topics/bbp/universal.html) when handling samples and controls.
n Clean and disinfect all spills of specimens or reagents, and other potentially contaminated materials.
SOP 1: Processing and handling blood specimens — pre-analytical phase
Date of creation or revision: / Prepared by: / Reviewed by: / Approved by:Purpose
n To describe how blood samples should be processed and handled before analysis to maintain sample integrity.
Note: For handling of DBS samples, refer to “Family-Centered Care of HIV-Exposed and HIV-Infected Children in Low-Resource Settings SOP 2.6: DBS Collection Procedure.” For handling of sputum, refer to “Implementation of TB Activities at HIV Service Delivery Sites Appendix 6: Visual representation of sputum collection procedure.”
Responsibility
n All individuals who collect, process, and handle samples, including but not limited to:
- Laboratory technicians
- Laboratory managers
- Physician or clinical officer
- Nurse or nurse assistant
- Couriers
Supplies
1. Gloves
2. Vacutainers (Becton, Dickinson and Company or equivalent)
3. Vacutainer storage racks
4. Equipment:
a. Centrifuge
b. Fridge
Procedure
1.1 Collect blood for serology, hematology, CD4 and biochemistry
n Refer to the Becton Dickinson and Company tube guide (Table 1, below) to determine the appropriate type of tube for the test requested. Refer to the SOP for the particular test requested to confirm which tube should be used.
n Collect blood in the following order (see Figure 2) to avoid contamination (if blood culture is required, collect sample for blood culture first). Reference site-specific venipuncture protocols for proper blood collection technique. Invert tubes the required amount (see Table 1) to ensure proper mixing of blood and additive. Ensure that all specimens are labeled correctly immediately following collection.
Figure 2: Order of blood collection by standard color of tube stopper
n Take precautions to avoid the following problems that can alter or inhibit analysis:
- Hemolysis
Do not keep a tourniquet on the patient’s arm for an extended period of time
Use a 20-gauge needle (larger gauge may be used for adults only)
Avoid expelling blood into a tube through a needle or applying excessive pressure on the plunger
Do not shake blood; always gently invert tubes
Do not refrigerate blood before it has clotted
- Bacterial changes/contamination
Use sterile technique; change gloves often
Process samples as soon as possible (see “1.2 Serum, plasma and whole blood processing”, below)
Store serum or plasma in fridge or freezer until analysis (take care to avoid multiple freeze-thaw cycles)
- Concentration changes
Do not rinse syringes or tubes with saline or anticoagulant solutions
Do not allow blood to stand in open containers before centrifugation
Process samples as soon as possible (see “Serum, plasma and whole blood processing”, below)
1.2 Process serum, plasma and whole blood
Before processing samples, refer to the product insert for tubes and/or test and review manufacturer’s recommendations and/or additional instructions.
n Process serum and plasma as follows:
- For Serum
Allow tube of blood to clot for 30 minutes at room temperature in vertical position
- Centrifuge for 10 minutes at 1300g. Tubes should be centrifuged as soon as possible and within 2 hours of collection.
- For Plasma
n Centrifuge for 10 minutes at 1300g. Tubes should be centrifuged as soon as possible and within 1 hour of collection. Visually inspect specimens to check sample integrity
- After centrifugation, serum/plasma should be clear, with a yellow tint, and should sit as the top layer (with red blood cells as the bottom layer). A cloudy top layer or red top layer (no clear separation) are indications that the sample may not be suitable for analysis
n Process whole blood as follows:
- Invert tube the required number of times to avoid clotting (Refer to Table 1: BD vacutainer® venous blood collection tube guide)
- No centrifugation is necessary
1.3 Handle and store sample
n Avoid excessive exposure to heat and light, and avoid freeze-thaw cycles.
n Take care to handle samples gently. Avoid shaking.
n Serum and plasma samples:
- Store serum and plasma at 2–8°C if analysis will be conducted within 7 days. If analysis is not expected to be conducted within 7 days, samples should be frozen at -20°C
n Whole blood samples (except for CD4 testing):
- Store whole blood at 2–8°C if analysis will be carried out within 7 days. DO NOT freeze whole blood. A new sample should be collected analysis does not take place within 7 days from the date of collection.
n Whole Blood Samples for CD4 testing:
- CD4 samples SHOULD NOT be refrigerated. Store CD4 samples at room temperature. Process as soon as possible and within time range specified by the test manufacturer and site specific SOP for CD4 testing, A new sample should be collected if analysis does not take place within the specified time range.
Table 1: BD vacutainer® venous blood collection tube guide
BD vacutainer® tubes with BD hemogard™ closure / BD vacutainer® tubes with conventional stopper / Additive / Inversions at blood collection* / Laboratory use /Full draw chemistry tubes
Gold / Red / Gray / n Clot activator and gel for serum separation / 5 / For serum determinations in chemistry. May be used for routine blood donor screening and diagnostic testing of serum for infectious disease**. Tube inversions ensure mixing of clot activator with blood. Blood clotting time: 30 minutes.
Light
Green / Green / Gray / n Lithium heparin and gel for plasma separation / 8 / For plasma determinations in chemistry. Tube inversions ensure mixing of anticoagulant (heparin) with blood to prevent clotting.
Red / Red / n Silicone coated (glass)
n Clot activator, silicone coated (plastic) / 0
5 / For serum determinations in chemistry. May be used for routine blood donor screening and diagnostic testing of serum for infectious disease**. Tube inversions ensure mixing of clot activator with blood. Blood clotting time: 60 minutes.
Orange / n Thrombin-based clot activator with gel for serum separation / 5 to 6 / For stat serum determinations in chemistry. Tube inversions ensure mixing of clot activator with blood. Blood clotting time: 5 minutes.
Orange / n Thrombin-based clot activator / 8 / For stat serum determinations in chemistry. Tube inversions ensure mixing of clot activator with blood. Blood clotting time: 5 minutes.
Royal Blue / n Clot activator (plastic serum)
n K2EDTA (plastic) / 8
8 / For trace-element, toxicology, and nutritional-chemistry determinations. Special stopper formulation provides low levels of trace elements (see package insert). Tube inversions ensure mixing of either clot activator or anticoagulant (EDTA) with blood.
Green / Green / n Sodium heparin
n Lithium heparin / 8
8 / For plasma determinations in chemistry. Tube inversions ensure mixing of anticoagulant (heparin) with blood to prevent clotting.
Gray / Gray / n Potassium oxalate / sodium fluoride
n Sodium fluoride / Na2 EDTA
n Sodium fluoride (serum tube) / 8
8
8 / For glucose determinations. Oxalate and EDTA anticoagulants will give plasma samples. Sodium fluoride is the anti-glycolytic agent. Tube inversions ensure proper mixing of additive with blood.
Tan / n K2EDTA (plastic) / 8
Partial-draw tubes (2ml and 3ml: 13 x 75mm)
Small-volume pediatric tubes (2ml: 10.25 x 47mm; 3ml: 10.25 x 64mm)
BD vacutainer® tubes with BD hemogard™ closure / BD vacutainer® tubes with conventional stopper / Additive / Inversions at blood collection* / Laboratory use
Partial draw and pediatric chemistry tubes
Red / n None / 0 / For serum determinations in chemistry. May be used for routine blood donor screening, immunohematology testing***, and diagnostic testing of serum for infectious disease**. Tube inversions ensure mixing of clot activator with blood. Blood clotting time: 60 minutes.
Green / n Sodium heparin
n Lithium heparin / 8
8 / For plasma determinations in chemistry. Tube inversions prevent clotting.
BD vacutainer® tubes with BD hemogard™ closure / BD vacutainer® tubes with conventional stopper / Additive / Inversions at blood collection* / Laboratory use
Full draw hematology and CD4 tubes
Lavender / n Liquid K3EDTA (glass)
n Spray-coated K2EDTA (plastic) / 8
8 / K2EDTA and K3EDTA for whole blood hematology determinations. K2EDTA may be used for routine immunohematology testing and blood donor screening***. Tube inversions prevent clotting.
Partial-draw tubes (2ml and 3ml: 13 x 75mm)
Small-volume pediatric tubes (2ml: 10.25 x 47mm; 3ml: 10.25 x 64mm)
BD vacutainer® tubes with BD hemogard™ closure / BD vacutainer® tubes with conventional stopper / Additive / Inversions at blood collection* / Laboratory use
Partial draw and pediatric hematology and CD4 tubes
Lavender / n Spray-coated K2EDTA (plastic) / 8
8 / For whole blood hematology determinations. May be used for routine immunohematology testing and blood donor screening***. Tube inversions prevent clotting.
* Invert gently, do not shake