Manual Microscopic Urinalysis Exam
Your Lab Name
Urinalysis
U.2. Manual Urinalysis Microscopic Exam
Manual Urinalysis Microscopic Exam
Author: Penny Stevens
Document Number:
Pro 68-0.1
Effective (or Post) Date:
9 June 2008
Document Origin
US Army Hospital Heidelberg, Germany
Company:
N/A
SMILE Approved by:
Heidi
SMILE Comments: This document is provided as an example only. It must be revised to accurately reflect your lab’s specific processes and/or specific protocol requirements. Users are directed to countercheck facts when considering their use in other applications. If you have any questions contact SMILE.
Copy # _____ Effective Date: Date
U. URINALYSIS
U.02. MANUAL URINALYSIS MICROSCOPIC EXAM
U.2.1. PRINCIPLE:
1. Routine Urinalysis consists of both physical and chemical analyses to assist physicians in the diagnosis and treatment of renal and urinary tract diseases and in the detection of metabolic or systemic disease processes not directly related to the kidney.
2. The microscopic examination of the centrifuged urine sediment includes the study of formed elements, such as WBC’s, RBC’s, casts and crystals.
3. The macroscopic examination of urine includes physical appearance, such as color, character and clarity. See U.1 Manual Urinalysis Dipstick SOP for macroscopic testing and reporting procedures.
4. A qualitative chemical analysis of the urine is performed by using a multi-parameter test strip that measure pH, protein, glucose, ketones, bilirubin, urobilinogen, nitrite, blood, leukocyte esterase, and specific gravity. The test strips are dipped in the urine and read visually according to the color comparison chart printed on the side of the container at prescribed time intervals. See U.1 Manual Urinalysis Dipstick SOP for qualitative testing and reporting procedures.
U.2.2.PURPOSE - The microscopic examination of urine sediment.
U.2.3. SPECIMENS
1. Use fresh well-mixed urine collected by clean-catch method into a sterile container.
2. The specimen should be unpreserved and uncentrifuged.
3. All urine specimens should reach the laboratory within one (1) hour after collection and be properly labeled.
4. Urine specimens should be tested within two (2) hours after collection. If urine cannot be tested within two (2) hours, it may be stored for up to four (4) hours at 2 to 8(C. (The specimen must be brought to room temperature before testing.)
5. The following urine samples are not satisfactory for testing:
5.1. Specimens received over two hours after collection.
5.2. Mislabeled samples.
5.3. Improperly collected samples. For example, urine samples with preservatives, specimens collected in non-sterile containers, or specimens collected in containers with soap or detergent residues will not be accepted.
5.4. QNS (Quantity Not Sufficient) - The recommended minimum volume is 12 mL’s. The required minimum volume for microscopic examination is 0.50 mL. In the event that less than <0.50 mL is received, perform testing on uncentrifuged urine and comment as noted in section U.2.7.
5.5. In the event that an unacceptable sample is received, another sample must be requested. Document all action taken in the LIS.
U.2.4.EQUIPMENT & REAGENTS
1. Equipment:
1.1 Kova System - Kova slides, pipettes & tubes
1.2. Centrifuge with timer
1.3. Microscope with 10x and 40x objectives
1.4. Lens paper or lint free tissues
1.5. Log sheets
1.6. Laboratory markers and pens
1.7. Clean gauze (4 x 4)
2. Reagents - None
U.2.5.CALIBRATION: Not applicable
U.2.6.QUALITY CONTROL:
1. Control Materials and Procedural Notes:
1.1. A Run in urinalysis is defined as all patient testing performed in a 24-hour period. Quality Control must be performed under the following conditions.
1.1.1. Every morning prior to testing patient specimens.
1.1.2. Whenever test results appear questionable.
1.2. A Normal and Abnormal Kova Control will be performed at least once every run.(see above)
1.3. Reconstitute new Kova controls with 60 mL Reagent Grade water according to manufacturer’s instructions.
1.3.1. After reconstitution, separate controls into 2 mL aliquots using disposable transfer pipettes and plastic test tubes. Cap tightly!
1.3.2. Label each plastic test tube with Normal or Abnormal Kova Control, lot number, expiration date, reconstitution date, and initials of tech who reconstitutes controls.
1.3.3. Sample aliquots will be stored frozen at -20(C. The sample aliquots are stable at this temperature for up to four (4) months.
1.3.4. Only one freeze/thaw cycle is permitted. Aliquots must be at room temperature before testing. Do not use a warming block to thaw.
1.3.5. Sample aliquots are stable for one hour at 15-20°C or 24 hours at 2-8°C.
1.4. Perform the Kova controls in the same manner as patient samples. See section U.2.7 for the patient testing procedure. Record the results on the Urinalysis Quality Control Worksheets. See appendix 3 & 4.
1.5. All results must fall into the expected ranges, which are obtained from the manufacturer’s insert for each control and recorded on the top of the Urinalysis Quality Control Worksheets.
1.6. If all results are within expected ranges, proceed with patient testing.
1.7. If QC results are outside of expected ranges for controls:
1.7.1.Re-examine the specimen using a different field of view.
1.7.2.Adjust the coarse/fine focus as needed to improve the view and repeat testing.
1.7.3.Check for signs of contamination in the controls.
1.7.4.Repeat the procedure with freshly reconstituted Kova controls.
1.7.5.If the results are still out of range after performing the above steps, notify the next higher supervisor immediately. Corrective action must be taken and QC must be in range before patient testing can be performed.
1.7.7.Record all QC values (both in and out of range) and corrective actions on the Quality Control Worksheets, appendix 3 & 4.
1.8.Parallel Testing: All new quality control lots or separate shipments of current lots will be tested against the current lots for performance verification before performing patient testing. See the Parallel Testing form, Appendix 5. Results are considered acceptable if they are within their lot number quality control limits.
U.2.7.PROCEDURE
1. Remove the specimens/controls from the refrigerator or freezer, as applicable, and allow them come to room temperature for at least 30 minutes before mixing.
2. Positively identify the specimens using the patient name and date of birth.
3. Log the sample on the testing worksheet using the LIS generated patient label, appendix 6.
4. Add the LIS generated label to the sample cup (not the sample lid). Any discrepancy must be investigated before processing the specimen.
5. Mix patient urine sample by swirling. Mix control aliquots by inverting several times to ensure homogeneity of the contents.
6. If a macroscopic examination & urine chemistry testing is ordered refer to U.01.1 Manual Urinalysis Dipstick Testing SOP for testing and reporting procedures. The microscopic examination is performed after dipstick testing unless the provider requests otherwise.
7. If Clintest, Acetest or Ictotest is required as listed below or requested by the provider, perform testing on uncentrifuged urine prior to performing the microscopic exam. Refer to individual confirmatory test SOPs as listed in the references.
8. The Clinitest confirmatory test is required for all pediatric urine samples with patients age 2 years old or younger, regardless of whether it was requested or not. See Clinitest SOP U06.1.
9. Backup tests and/or microscopic examination must be performed on urine that tests positive for the following:
Positive Result
Backup Test
Microscopic Exam
Blood
N/A
Yes
Nitrite
N/A
Yes
Ketones
Acetest
No
Glucose
Clinitest
No
Bilirubin
Ictotest
No
Protein
3% Sulfosalicylic Acid
Yes
Leukocyte Esterase
N/A
Yes
10. Confirmatory tests are not required for TRACE results on PROTEIN or KETONES.
11.Microscopic examinations are required on the following specimens regardless of the reagent strip results:
11.1. Urine with cloudy or hazy appearance.
11.2.Microscopic examination requested by the patient’s physician.
11.3.Urine specimens received from Pediatric Clinic.
11.4.Catheterized and Suprapubic aspiration samples.
11.5.Urine specimens received from Physical Exam Clinic.
12. Pour 12-mL’s of specimen into a sterile Kova tube and label the tube with the LIS specimen label. If the total specimen volume is 0.50 mL or less, do not pour the sample into a Kova tube. Perform testing from the specimen cup as indicated in the following steps.
13. Testing
13.1. Volumes greater than 0.50 mL
13.1.1. Perform Dipstick, Clinitest, Acetest, or Ictotest as required or requested.
13.1.2.Cap the Kova sample and centrifuge at 1500 rpm for 5 minutes.
13.1.3.Remove the sample from the centrifuge and discard the cap.
13.1.4. If 3% SSA testing is required or requested, perform testing on the supernatant according to the U.10.1. 3% SSA SOP.
13.1.5. Decant and discard the supernatant while retaining the sediment pellet in the conical portion of the Kova tube. There will be a small volume of fluid retained above the sediment. The total volume retained in the Kova system tube after decanting the supernatant should be approximately 0.5 mL.
13.1.6.Use a Kova disposable pipette to resuspend the sediment pellet.
13.1.7. Proceed with testing in step 13.3.
13.2. Volumes less than 0.50 mL - swirl the sample in the specimen cup to mix well.
13.3. Use a Kova pipette to aspirate sample from the well mixed specimen.
13.4. Place one (1) drop of the sediment on the Kova slide so that the urine sediment is drawn under the preformed Kova cover slip.
13.5. Place the Kova slide under the microscope with the low power objective (10X lens) and observe the sediment for the presence of casts. Observe at least ten (10) fields and take the average number of casts seen for each type.
13.6. Switch to high power objective (40X lens) to observe the sediment for the presence of epithelial cells, WBCs, RBCs, bacteria, crystals, yeast, trichomonas, mucus, and spermatozoa.
13.7. Scan approximately 8-10 fields and take the average per field for each type of sediment.
13.8. QNS specimens - report the following canned comments as applicable:
13.8.1. If the volume was <12 mL but > 0.5 mL, enter [12ML] - Minimum specimen volume is 12 mL. Urine sediment results may be falsely decreased due to a submitted sample volume less than 12 mL’s.
13.8.2. If the volume was <0.5 mL, enter [UNSPUN] - Testing performed on uncentrifuged urine due to a submitted volume less than 0.5 mL. Minimum required specimen volume is 12 mL. Urine sediment results may be falsely decreased.
13.9. Review the results and enter data on the manual microscopic patient result form (appendix IV). See section U.2.8 for Expected and Critical Results - Abnormal results will be flagged.
13.10. Enter all results in the LIS. Do not certify any results until all urinalysis testing ordered on the patient sample is complete.
13.11. Urine samples are retained at 2-10°C for 24 hours in the Hematology refrigerator.
U.2.8. EXPECTED / CRITICAL RESULTS & REPORTABLE RANGE:
1. The following bolded analytes require a result entry during the microscopic results even if they are not seen during the microscopic exam. Report “none” or 0-1 or 0-2 as indicated. All other analytes are optional entry if seen during the microscopic exam.
Analyte
Expected results for: ALL AGES
Reportable Range:
HYALINE CASTS
0-1
0-1, 1-3, 3-5, 5-10, 10-25, 25-50, or greater than (>) 50/ LPF. Quantify each cast type separately
Granular Cast
None
Cellular Cast (RBC, WBC, etc)
None
Waxy Cast
None
EPITHELIAL CELLS
0-2
0-2, 2-5, 5-10, 10-25, 25-50, or greater than (>) 50/ HPF
LEUKOCYTES (WHITE BLOOD CELLS)
0-2
0-2, 2-5, 5-10, 10-25, 25-50, or greater than (>) 50/ HPF
RED BLOOD CELLS
0-2
0-2, 2-5, 5-10, 10-25, 25-50, or greater than (>) 50/ HPF
BACTERIA
None - Few
None, Few, Trace, Moderate or Many
MUCUS
None - Light
Light, Moderate or Heavy
Crystals
None
Few, Moderate or Many for each crystal type.
Spermatozoa
Males only: Few
Few, Moderate, or Many.
Yeast
None
Light, Moderate or Many. Report any budding yeast or hyphea seen using comments as noted below.
Trichomonas
None
Few, Moderate or Many. May only be reported if motile.
2. If budding yeast or hyphea are present, add the following canned comments:
2.1. [Budding] - Budding yeast present.
2.2. [Hyphea] - Hyphae present.
3. The presence of sperm in the urine of a female child under 16 years old is a critical value. Supervisor confirmation is required before reporting any suspected sperm to the physician. The physician must be notified in accordance with critical value procedures before sperm is reported in the LIS. The urine must be retained at 2-10°C until the patient’s physician and laboratory director authorizes disposal. In no instance will the urine be destroyed until duly authorized.
4. If a urine dipstick is performed, compare the results obtained on the microscopic with the multi-parameter reagent strip with the following facts in mind:
4.1.Urine with RBCs seen on the microscopic exam should have a positive occult blood on the reagent strip.
4.2.Urine with casts should have elevated specific gravity and positive protein.
4.3.Urine with crystals should have elevated specific gravity.
4.4.Urine with positive nitrite should have bacteria on the microscopic.
5. CRITICAL VALUES:
5.1. When the following values are encountered after a urinalysis testing, immediately notify the physician or senior ward/clinic nurse of the values encountered.
Analyte
Critical Results: All ages unless otherwise noted
Waxy Casts
Any
Red Blood Cell Cast
Any
Cystine Crystals
Any
Tyrosine Crystals
Any
Leucine Crystals
Any
Sperm
Any - Females < 16yrs old
5.2. Critical values must be reported in accordance with the laboratory critical value policy.
5.2.1. All critical microscopic sediment must be verified by the laboratory supervisor before reporting to the physician/nurse and before certifying results in the LIS.
5.2.2.All results must be read back by the physician/nurse and the notification & read back documented in the LIS.
5.2.3.The pathologist must also be notified within 2 hours and will determine if a pathologist review is required.
U.2.9. PROCEDURAL NOTES:
1. NORMAL CHARACTERISTICS OF THE URINE: The yellow color of the urine is due largely to the pigment urochrome and small amounts of urobilin and uroerythrin. Normal urine is essentially clear, and the presence of particulate matter in uncentrifuged urine needs to be explained microscopically. Normal urine has a faint, aromatic odor of undetermined source.
2.Microscopic Sediment:
2.1. RBCs: Normally 0-2 RBCs/HPF may be seen in urine from males and non-menstruating females. Increased numbers may indicate renal hematuria.
2.2. WBCs: Normally 0-5 WBCs/HPF may be seen in urine of normal males with slightly higher ranges in females. Increased numbers may indicate renal disease or acute infection.
2.3. Epithelial cells: A few epithelial cells are normal and indicate normal sloughing off of aging cells. Increased numbers may indicate renal disease, urinary tract infection, or poor technique in specimen collection.
2.4. Casts: 0-1 hyaline cast/LPF is found in normal urine. Increased numbers or more advanced types indicate proteinuria.
2.5. Bacteria: A few bacteria are normally seen due to poor technique in collection of the specimen. Increased numbers may indicate kidney, bladder, or urinary tract infection.
2.6. Crystals: The following crystals may be seen in normal or abnormal urine as noted below. Use urine pH and solubility information (appendix 7) to aid in identification, as needed. Also use appropriate image and literature resources to assist with identification.
Normal
Abnormal
Acidic Urine
Alkaline Urine
Acidic Urine
Alkaline Urine
Amorphous Urates
Amorphous Phosphates
Cystine
None
Uric Acid
Triple Phosphates
Tyrosine
Calcium Oxalate
Ammonium Biurates
Leucine
Calcium Phosphates
Sulfonamide (Sulfadiazine)
Calcium Carbonates
2.7. Mucus: Light mucus is normally found in urine and can sometimes be confused with hyaline casts.
2.8. Spermatozoa: A few are normally found in urine from normal males. Increased numbers are found in prostatic disease.
2.9. Trichomonas: Normally not found in urine. Presence of trichomonas is abnormal and may indicate infection.
5.10.Yeast: 0-1/HPF or few per high power field are normal in females. Increased numbers indicates infection.
5.11.Casts are classified according to their matrix, inclusions, pigments and cells present.
5.11.1.Cast matrices:
5.11.1.1.Hyaline casts: Are translucent cylindrical structures composed of mucoprotein. Increased numbers are seen with renal diseases and transiently with exercise, fever, congestive heart failure, and diuretic therapy.
5.11.1.2.Waxy casts: These differ from hyaline casts in that they are easily visualized because of their high refractive index. Waxy casts are homogeneously smooth in appearance. Their margins are sharp, their ends are blunt, and cracks or convolutions are frequently seen along the lateral margins. Waxy casts are commonly associated with tubular inflammation and chronic renal failure. They are also found during acute or chronic renal allograft rejection.
5.11.2.Cast Inclusions:
5.11.2.1.Granular casts are semitransparent cylinders containing small (fine) or large (coarse) granules. These granules represent plasma protein aggregates. Granular casts appear with glomerular or tubular diseases.
5.11.2.2.Fatty casts: Are semitransparent or granular cylinders containing large highly refractive vacuoles or droplets. Visible fat droplets are triglycerides or cholesterol esters. These are commonly seen when there is heavy proteinuria and are a feature of the nephrotic syndrome.
5.11.2.3.Crystal Casts: Crystalline inclusion in a semitransparent or granular cylinder. These casts indicate disposition of crystals in the tubule or collecting duct.
5.11.3.Cast Pigments:
5.11.3.1.Hemoglobin (Blood) casts: Appears yellow to red; sometimes the color is very pale and difficult to interpret. These casts are associated with glomerular disease.
5.11.3.2.Myoglobin casts: These cast are red-brown in color and occur with myoglobinuria following acute muscle damage.
5.11.3.3.Bilirubin and other drug casts: Bilirubin is seen in urine when there is obstructive jaundice, and will color casts as deep yellow brown. Drugs such as phenazopyridine (Pyridium) cause a bright yellow to orange color in acid urine and will color casts and cells.
5.11.4.Cellular Casts:
5.11.4.1.Erythrocyte (Red Blood Cell) casts: Semitransparent or granular cylinders containing distinct erythrocyte. Disorders reflected in the presence of erythrocyte casts in the sediment may include acute glomerulonephritis, IgA nephropathy, lupus nephritis, subacute bacterial endocarditis, and renal infarction.
5.11.4.2.Leukocyte (White Blood Cell) Casts: Semitransparent or granular cylinders containing leukocytes. They may be seen pyelonephritis, glomerular diseases, interstitial nephritis, lupus nephritis, and nephrotic syndrome.
6.SOURCES OF ERROR:
6.1. Urine should be tested within two (2) hours after collection. Prolonged testing delay may result in cast dissolution, RBC crenation or bursting, increased bacteria, and crystals dissolution.
6.2. Fill the Kova slide chamber with the pipette placed parallel to the slide and dispense the specimen drop in the corner of the well. Samples placed in the center of the well can cause air bubbles to form resulting in sample distortion during examination.
U.2.10. APPENDICES:
1. SOP Validation Form and SOP Change Control
2. SOP Approval
3. Urinalysis Normal Quality Control Worksheet
4. Urinalysis Abnormal Quality Control Worksheet
5. Quality Control Parallel Testing Worksheet
6. Patient Result Form
7. Urine Crystal Properties
U.2.11. REFERENCES:
1. Stransinger, Susan K., Urinalysis and Body Fluids, Third Edition, F.A. Davis Book Publisher, 1994, Pages 1 to 10 and 51 to 74.
2. Haber, Meryl H., Urinary Sediment: A Textbook Atlas, American Society of Clinical Pathologist Book Publisher, 1994.
3. Multistix 10 with SG Package Insert, Bayer Corporation; Diagnostics Division, 1999.
4. Kova Trol: Human Urinalysis Controls Package Insert, Hycor Biomedical Inc., 2001.
5. Manual Urinalysis by Bayer 10-SG Multistix SOP, U.1.1
6. Specific Gravity Determinations by Refractometer SOP, U.5.1
7. Clinitest Determination of Reducing Substances in Urine SOP, U.6.1
8. Acetest Determination of Ketones in Urine SOP, U.8.1
9. Ictotest Determination of Bilirubin in Urine SOP, U.9.1
10. SSA Determination of Protein in Urine SOP, U.10.1
Appendix 1
SOP VALIDATION
SOP NAME:
U.2. Manual Urinalysis Microscopic Exam
Clear and specific title and principle: yes / no
Comments:
All necessary supplies, equipment, and materials are listed: yes / no
Comments:
SOP is sufficiently detailed to be understood but not overly complex: yes / no
Comments:
SOP text adequately describes process/procedure: yes / no
Comments:
SOP accomplishes purpose: yes / no
Comments:
Reviewed by: (Name & Title) )
Signature: __________________ Date: __________________
SOP CHANGE CONTROL
Date Change QA OIC Med. Dir.
Appendix 2
SOP APPROVAL
SIGNATURE
DATE
PREPARER
QA COORDINATOR
LABORATORY OIC
MEDICAL DIRECTOR
ANNUAL REVIEW
REVIEWER SIGNATURE DATE REVIEWER SIGNATURE DATE
DOCUMENT COPY CONTROL
DATE:___________ # COPIES __________
LOCATIONS
SUPERSEDES:
DATE SOP RETIRED: __________
Appendix 3
Urinalysis Dipstick and Microscopic Normal Control Log
Normal Control Lot#:
Expiration:
Month/Year:
Dipstick Lot#
Expiration:
Date in use:
Supervisor Review:
Dipstick Lot#
Expiration:
Date in use:
Review Date:
Color/ Clarity
Glucose
Bilirubin
Ketones
Specific Gravity
Blood
pH
Protein
Urobilin.
Nitrate
Leu. Esterase
WBC/hpf
RBC/hpf
Crystals
Casts
Tech Initials
Date
Acceptable Range:
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
Comments:
Refer to SOPs U.1.1 and U.2.1 for Urine Dipstick and Microscopic QC resulting procedures
Appendix 4
Urinalysis Dipstick and Microscopic Abnormal Control Log
Abnormal Control Lot#:
Expiration:
Month/Year:
Dipstick Lot#
Expiration:
Date in use:
Supervisor Review:
Dipstick Lot#
Expiration:
Date in use:
Review Date:
Color/ Clarity
Glucose
Bilirubin
Ketones
Specific Gravity
Blood
pH
Protein
Urobilin.
Nitrate
Leu. Esterase
WBC/hpf
RBC/hpf
Crystals
Casts
Tech Initials
Date
Acceptable Range:
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
Comments:
Refer to SOPs U.1.1 and U.2.1 for Urine Dipstick and Microscopic QC resulting procedures
Appendix 5
Quality Control Parallel Testing Worksheet
Reagent
Current Lot Result
Current Lot Acceptable Range
New Lot Result
Current Lot Acceptable Range
Acceptable
Red Blood Cells
Yes / No
White Blood Cells
Yes / No
Casts
Yes / No
Crystals
Yes / No
*Bacteria
None
None
Yes / No
Yes / No
Yes / No
Yes / No
Yes / No
Yes / No
*Note: If bacterial presence is suspected in the quality control material, it is unacceptable for use. Notify the supervisor immediately.
New Lot Acceptable:
Yes / No
Comments:
Tech Signature:
Date:
Supervisor review :
Date:
Supervisor Comments:
Appendix 6 - Patient Manual Urinalysis Result Report
Analyte
Patient Results
Reference Range
Hyaline Casts / LPF:
0-1
1-3
3-5
5-10
10-25
25-50
> 50
0-1 / LPF
Other Casts:
None / LPF
Type:
Type:
Epithelial Cells /HPF
0-2
2-5
5-10
10-25
25-50
> 50
0-2 / HPF
Leukocytes (WBC’s) / HPF
0-2
2-5
5-10
10-25
25-50
> 50
0-2 / HPF
Red Blood Cells (RBC’s) / HPF
0-2
2-5
5-10
10-25
25-50
> 50
0-2 / HPF
Bacteria / HPF
None
Few
Trace
Moderate
Many
None-Few / HPF
Mucus / HPF
None
Light
Moderate
Heavy
None-Light / HPF
Crystals / HPF:
None
Few
Moderate
Many
None / HPF
Type:
None / HPF
Type:
None / HPF
Spermatozoa / HPF:
None
Few
Moderate
Many
Males Only: Few / HPF
Yeast / HPF:
None
Few
Moderate
Many
None / HPF
Trichomonas
None
Few
Moderate
Many
None / HPF
Comments:
Tech Signature:
Report Date/Time:
Supervisor review required for all critical values. Required?
Yes / No
Signature:
Date:
Comments:
Appendix 7 - Urine Crystal Properties
Technician: _______________________________ Date:___________________________________
New Control Lot#: __________________________ Exp. Date: _________________________________
Current Control Lot#: _________________________ Exp. Date: _________________________________
Sample Information:
Date Collected: ____________________________
Time Collected: ____________________________
Ordering Physician & Clinic: _____________________
Patient Information:
Patient Name: _____________________________
ID# ___________________________________
Date of Birth: ______________________________
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