SOP-H-004 — Erythrocyte Sedimentation Rate (ESR — Modified Westergren Method) SOP
Type: SOP · Version: V0 · Status: DRAFT — UNAPPROVED · Served read-only from the bundled QMS copies. The Drive master is untouched.
Rendered in-browser from the project's Word file SOP-H-004-erythrocyte-sedimentation-rate-westergren-method-sop.docx (policies-docx / forms-docx — the file Junry reviews). Formatting is faithful, not pixel-perfect; the DOCX is the official copy.
1.0 Purpose
This SOP sets the procedure for performing the Erythrocyte Sedimentation Rate (ESR) by the Modified Westergren Method at Tarangnan Community Hospital. It covers specimen handling, dilution requirements, strict vertical tube leveling, timing, reference intervals, and technical interference controls for measuring non-specific systemic inflammation.
2.0 Scope
This procedure applies to all Medical Technologists performing manual sedimentation testing and Laboratory Aides providing workbench support in the Hematology Section. It covers manual and semi-closed Westergren sedimentation testing of pediatric and adult outpatient and inpatient specimens.
3.0 Definitions
- Erythrocyte Sedimentation Rate (ESR): The rate at which anticoagulated red blood cells settle vertically in a standardized column of plasma over a defined one-hour period, expressed in millimeters per hour (mm/hr).
- Rouleaux Formation: The reversible face-to-face aggregation of red blood cells resembling stacks of coins, facilitated by asymmetric, positively charged plasma proteins (fibrinogen, immunoglobulins) that reduce erythrocyte repulsive zeta potential.
- Modified Westergren Method: The reference sedimentation technique approved by the International Council for Standardization in Haematology (ICSH) using K2EDTA whole blood diluted at a 4:1 volumetric ratio with trisodium citrate solution in a calibrated 200 mm vertical tube.
- Buffy Coat: The thin layer of packed leukocytes and platelets situated immediately above the settled red cell column; excluded when reading the sedimentation boundary.
4.0 Responsibility
4.1 Hospital Director
- Approves this SOP and provides the laboratory infrastructure, including level, vibration-free countertops and certified Westergren testing apparatus.
4.2 Pathologist
- Reviews and authorizes reference intervals, clinical utility guidelines, and quality assurance protocols for ESR testing.
- Advises attending physicians on the diagnostic interpretation of extreme or clinically discordant sedimentation rates.
4.3 Chief Medical Technologist / Laboratory Quality Manager
- Ensures technical compliance with this SOP, verifies annual leveling of ESR racks, and monitors commercial QC or duplicate testing records.
- Oversees reagent quality, timer calibration, and technologist competency evaluations.
4.4 Medical Technologists
- Inspect specimens for volume and clot integrity, prepare standardized 4:1 blood-to-citrate dilutions, fill Westergren pipettes, and position tubes in calibrated vertical racks.
- Maintain exact 60-minute timing, read the sedimentation meniscus excluding the buffy coat, and record results on the TCH Hematology Result Form.
- Identify and document technical sources of error, including temperature fluctuations, rack vibration, and bubble formation.
4.5 Laboratory Aides
- Clean and decontaminate ESR racks and surrounding work surfaces at the conclusion of each shift.
- Transport designated ESR specimens from the accessioning counter to the hematology workstation.
- Shall never fill Westergren tubes, set timers, read sedimentation levels, or record ESR results.
5.0 Policy / Procedure
5.1 Principle of the Modified Westergren Method
- When anticoagulated whole blood is placed into a vertical, calibrated tube, erythrocytes sediment downward under gravity while plasma rises.
- The sedimentation process occurs in three distinct kinetic phases: - Aggregation Phase (First 10 minutes): Erythrocytes form microscopic rouleaux stacks. - Precipitation / Fast Settling Phase (Middle 40 minutes): Rouleaux aggregates sediment rapidly at a constant velocity. - Packing Phase (Final 10 minutes): Erythrocytes accumulate and pack tightly at the bottom of the column.
- The distance settled by the top of the red cell column in exactly 60 minutes reflects plasma protein composition, particularly acute-phase reactants (fibrinogen, alpha-2 macroglobulin, immunoglobulins) that neutralize negative red cell surface charges.
5.2 Specimen Requirements and Stability
- Acceptable Specimen: Whole blood collected into a dipotassium ethylenediaminetetraacetic acid (K2EDTA lavender-top) vacuum tube.
- Minimum Volume: 2.0 mL whole blood (1.0 mL for dedicated microtainer systems).
- Specimen Stability: - Specimens shall be tested within 4 hours of collection when stored at room temperature (18–25 °C). - If testing cannot be performed within 4 hours, the specimen may be refrigerated at 2–8 °C for up to 24 hours. - Refrigerated specimens must be allowed to warm to room temperature for at least 30 minutes and inverted 8 to 10 times before dilution.
- Specimen Rejection: - Grossly hemolyzed, clotted, or severely underfilled tubes (< 50% nominal volume) shall be rejected immediately. - Specimens older than 24 hours shall not be tested, as aging erythrocytes become spherical and resist rouleaux formation, causing falsely low ESR.
5.3 Equipment, Reagents, and Materials
- Westergren Pipettes: Standard glass or disposable plastic pipettes having an internal bore diameter of 2.55 mm ± 0.15 mm and a calibrated graduation scale from 0 to 200 mm.
- ESR Leveling Rack: Specialized leveling stand equipped with a built-in bubble spirit level, plumb line, spring-loaded clips, and leak-proof silicone gaskets.
- Diluent: Freshly prepared or commercial 3.8% (0.129 mol/L) or 3.2% (0.109 mol/L) trisodium citrate dihydrate solution.
- Calibrated Laboratory Timer: Digital timer accurate to within ±1 second.
- Micropipettes: Precision volumetric pipettes (800 µL and 200 µL, or 1.6 mL and 0.4 mL) with disposable tips.
- Closed Westergren Reservoirs: Single-use pre-filled citrate vials (e.g., Seditainer or equivalent closed aspiration systems) if used by the section.
5.4 Stepwise Testing Procedure
- Environmental Verification: Ensure the ESR leveling rack is positioned on a rigid, heavy workbench completely isolated from centrifuges, vortex mixers, mechanical rotators, direct sunlight, and drafts from air conditioning vents.
- Verify that the rack spirit level indicates a strictly horizontal alignment (0° deviation).
- Dilution Preparation (Manual Method): - Invert the K2EDTA patient tube 10 times to ensure uniform cell distribution. - Pipette 0.4 mL (400 µL) of 3.8% trisodium citrate diluent into a clean 12 × 75 mm glass or plastic tube. - Add 1.6 mL (1,600 µL) of well-mixed whole blood (maintaining an exact 4:1 blood-to-citrate ratio). - Cap the tube and invert gently 8 to 10 times to mix thoroughly. Avoid frothing.
- Filling the Westergren Column: - Draw or push the diluted blood into the Westergren pipette up to exactly the 0 mm graduation mark. - Ensure the blood column is completely free of trapped air bubbles or gaps. - Seat the pipette firmly into the vertical rack, ensuring the base is sealed against the lower rubber gasket and the top is held securely by the spring clamp.
- Incubation and Timing: - Immediately start the digital timer set for exactly 60 minutes (1 hour ± 1 minute). - Do not disturb, vibrate, touch, or move the rack or surrounding bench during the 60-minute settling period.
- Reading Results: - At exactly 60 minutes, read the distance from the bottom of the surface plasma meniscus to the top of the settled erythrocyte column. - The result is read in whole millimeters (mm). - Exclusion of Buffy Coat: If a white/grey buffy coat layer is visible above the red cell column, exclude it; read only to the upper margin of the packed red cell column. - Record the value as millimeters in the first hour (mm/hr).
5.5 Quality Control and Procedure Verification
- Commercial whole-blood ESR control material (bi-level Normal and Abnormal) shall be analyzed once weekly or with each new lot of Westergren tubes/diluent, where available.
- In the absence of commercial controls, the section shall conduct duplicate split-sample testing on one patient specimen daily across two separate Westergren racks. The two readings must agree within ±3 mm/hr (or within 10%, whichever is greater).
- If duplicate readings fail, investigate rack leveling, pipette bore defects, gasket leakage, or bench vibration.
- Document all QC and duplicate checks in the TCH Daily QC Log.
5.6 Reference Intervals
| Patient Population | Age Category | Reference Interval (mm/hr) |
|---|---|---|
| Adult Male | < 50 years | 0 to 15 mm/hr |
| Adult Male | ≥ 50 years | 0 to 20 mm/hr |
| Adult Female | < 50 years | 0 to 20 mm/hr |
| Adult Female | ≥ 50 years | 0 to 30 mm/hr |
| Pediatric | < 18 years | 0 to 10 mm/hr |
| Newborn | Birth to 1 month | 0 to 2 mm/hr |
5.7 Technical and Clinical Interferences
- Technical Factors Causing Falsely Elevated ESR: - Rack Incline / Tilting: A tube tilted as little as 3° from the vertical accelerates red cell settling by up to 30% due to plasma channeling along the upper wall. - Elevated Temperature: Ambient temperatures > 25 °C reduce plasma viscosity, accelerating sedimentation. - Bench Vibration: Vibration from centrifuges or vortex mixers on the same bench accelerates sedimentation. - Underdilution: Excess blood volume relative to citrate increases apparent settling.
- Technical Factors Causing Falsely Decreased ESR: - Specimen Delay (> 4 hours at RT): Red cells lose discocyte morphology and become spherical, impeding rouleaux formation. - Low Ambient Temperature: Room temperatures < 18 °C increase plasma viscosity, slowing sedimentation. - Overdilution: Excess citrate dilutes plasma proteins, impeding rouleaux. - Air Bubbles / Clots: Microclots or bubbles disrupt the continuous vertical fluidic column.
- Clinical Factors Influencing ESR: - Decreased ESR: Polycythemia vera, sickle cell anemia (abnormal morphology prevents stacking), spherocytosis, marked leukocytosis, hypofibrinogenemia, congestive heart failure. - Elevated ESR: Severe anemia (reduced cell volume accelerates plasma separation), multiple myeloma and macroglobulinemia (marked paraprotein rouleaux), systemic infections, autoimmune connective tissue disorders, acute myocardial infarction, pregnancy, and advanced malignancy. - In patients with extreme anemia (Hct < 20%), ESR is often elevated secondary to low cell mass and should be interpreted alongside C-reactive protein (CRP).
6.0 Records
| Record Description | Controlled Form / Log | Storage Location | Retention Authority |
|---|---|---|---|
| Released ESR Patient Test Reports | LIS Server / F-QM-POST-001-A-V0 | Medical Records Archive | Retained per Laboratory Retention Schedule Policy |
| Daily ESR Bench Worksheets | Hematology Section Logbook | Hematology Workstation | Retained per Laboratory Retention Schedule Policy |
| ESR Duplicate and Commercial QC Records | Hematology QC Binder | Quality Control Cabinet | Retained per Laboratory Retention Schedule Policy |
| Equipment Maintenance and Environmental Log | F-QM-ANA-005-A-V0 | Equipment Dossier | Retained per Laboratory Retention Schedule Policy |
| Daily Facility and Environment Safety Log | F-QM-SAF-002-A-V0 | Laboratory Safety File | Retained per Laboratory Retention Schedule Policy |
7.0 References
- DOH Clinical Laboratory QMS Tool, Section 3 — Service Delivery
- DOH Administrative Order No. 2021-0037, Annex B1 — Technical Standards for Clinical Laboratories
- ISO 15189:2022, Clause 7.3 Examination processes
- CLSI H02-A5, Procedures for the Erythrocyte Sedimentation Rate Test
- International Council for Standardization in Haematology (ICSH), Reference Method for the Erythrocyte Sedimentation Rate
Companion policies:
- Hematology Section Workflow Quality Control and Result Validation Sop — Hematology Section quality control, interference, and result validation SOP
- Laboratory Reporting Policy — Result review, validation, and diagnostic report release
- Laboratory Retention Schedule Policy — Retention schedules for hematology worksheets and patient reports