unit 2 Chemical Urinalysis Reagent Strips - Comprehensive Study Notes
Learning Objectives
Examine chemical urinalysis testing.
Describe the reagents used to test for various analytes on a chemical reagent strip.
Explain internal and external quality control testing for chemical urinalysis reagent strips (manual and automated).
Discuss the significance of specific gravity and other positive analytes.
Understand normal reference ranges for all chemical tests.
Correlate chemical urinalysis results to disease processes.
Understand normal and critical chemical urinalysis results and the follow-through by an MLT for verification.
Determine when to order a microscopic urinalysis testing based on chemical results.
Discuss urine specimen retention, storage, and disposal.
Reagent Strips: Advantages and Manufacturer Guidance
Reagent strips are advantageous for chemical urinalysis due to convenience, cost-effectiveness, stability, ease of learning, disposability, smaller sample volumes, and space savings.
Follow the manufacturer’s directions for all reagent strip tests.
Always read the product insert that comes with each container of commercial reagent strips.
Storage and General Precautions
Keep reagent strips in a tightly capped container, away from moisture, direct sunlight, hit or volatile substances.
Each container includes a desiccant to protect strips from moisture.
Store at room temperature, under 30°C.
Strips must remain in their original containers; do not mix strips from different containers.
Do not touch the test area; ensure strips are not expired; open when ready to test and replace the cap immediately after removal.
Stability and expiry are critical for accurate results.
Stability and Timing Considerations
Each container is labeled with a lot number and expiration date.
Use strips within the expiration date; once opened, use within six months.
Discard the entire bottle if any reagent pads are damaged/compromised or if quality control (QC) is consistently out of range.
Read results at the time specified by the manufacturer for each chemical test.
Note: Different results may be seen for the same specimen at different times.
Reading Results: Manual vs Automated Methods
Manual method:
1) Dip the strip into urine and start the timer.
2) Hold the test pad next to the ID band on the container.
3) Orient the strip correctly to the color chart per manufacturer instructions.
4) Match the closest color block for each chemical test at the proper time.
5) Record results on the worksheet.Automated or semi-automated readers eliminate human variation in color interpretation and improve reproducibility.
Examples of automated systems include the URYXXON Relax strip reader (Macherey-Nagel) and related models.
Quality Control (QC)
Use both negative and positive controls to test every parameter of the reagent strip in use.
Perform QC at least once per day or per shift, and whenever a new container of strips is opened.
Document QC results in a dedicated binder for lab records.
QC results should be compared against manufacturer-provided expected results for both negative and positive controls.
Note on interpretation: manufacturer-provided results are not patient reference ranges; they are for QC purposes.
QC Documentation Examples (from the provided materials)
Negative Control sample (example):
GLUCOSE: Negative
BILIRUBIN: Negative
KETONES: Negative
SPECIFIC GRAVITY: 1.015 (within a tested expectation of 1.010–1.035 range)
BLOOD: Negative
pH: 7.0
PROTEIN: Negative
UROBILINOGEN: 3.2 µmol/L
NITRITE: Negative
LEUKOCYTES: Negative
Date Tested: 2023/03/10 (example)
Note: Two consecutive negative QC results; indication that results look good according to the manufacturer.
Positive Control sample (example):
GLUCOSE: 14 mmol/L
BILIRUBIN: +++
KETONES: 8 mmol/L
SPECIFIC GRAVITY: 1.015
BLOOD: +++
pH: >9.0
PROTEIN: 1.0 g/L
UROBILINOGEN: 33 µmol/L
NITRITE: Positive
LEUKOCYTES: ++
Date Tested: 2023/04/10
Note: Two days of positive QC results; indicates the positive control was within range per the manufacturer.
Important note: These QC results are for manufacturer-provided controls and are not patient reference ranges.
QC Troubleshooting Guidance
If a value falls outside the expected range:
Circle the outlier (red is suggested to draw attention).
If only one level (e.g., Pos or Neg) is out of range, re-test that level.
First step: verify strips are not expired; repeat with the same QC reagent to check for random QC error.
If results return to within range, document the new value alongside the previous one with lot/expiry information.
If re-testing fails to produce expected results:
Check timing of the reading, replace with a new container of strips, or replace QC reagents.
Consider lot number issues, expired reagents, or other QC reagent problems; use fresh reagents with proper expiry.
Specimens for Testing: Specimen Requirements
Fresh or adequately preserved specimens (refrigerator or chemical preservatives).
If no preservative is used, test within 2 hours of collection.
Well mixed; do not centrifuge.
Test specimen at room temperature.
Manual Testing Procedure (Summary)
1) Mix urine by gently swirling.
2) Remove strip, recap bottle tightly.
3) Immerse strip in urine; remove immediately to avoid dissolving reagents.
4) Wipe edge of strip to remove excess urine.
5) Hold strip horizontally; align with color blocks on label at specified times.
6) Record results on the worksheet.
Test Strips and Analytes
Glucose, Bilirubin, Ketone, Specific Gravity, Blood, pH, Protein, Urobilinogen, Nitrite, Leukocytes
Normal Chemical Urine Properties (Reference Values)
Glucose: Negative
Ketone: Negative
Protein: Negative
Specific Gravity: 1.001 ext{ to } 1.035 ext{ (adult random urine)}
Blood: Negative
pH: 5.0 ext{ to } 8.0
Bilirubin: Negative
Urobilinogen: 3.2 ext{ to } 17.0 ext{ µmol/L}
Nitrite: Negative
Leukocytes: Negative
Note: Reference values are based on Anderson College CHEMISTRY REFERENCE VALUES and Siemens Multistix 10SG Product Insert; small variances may occur by clinical site.
Glucose: Clinical Significance
Glucosuria/glycosuria = glucose in urine.
Glycosuria may occur after large sugar intake or during pregnancy.
Pathological: glucose in urine is an indicator of diabetes mellitus.
Non-pathological: menstruation or strenuous exercise can cause temporary glucose in urine.
Renal threshold for glucose is approximately ext{threshold}_{ ext{glucose}} \approx 10 \, \mathrm{mmol/L}; when blood glucose exceeds this threshold, glucose appears in urine (glycosuria).
Glucose Testing Principle and Interferences
Principle: Glucose oxidase-based test; double-sequential enzyme reaction (glucose oxidase in step 1, peroxidase in step 2).
Sensitivity (minimum detectable level): 4.2 \, \text{to} \, 6.9 \, \mathrm{mmol/L}.
Historical reference: Clinitest (non-specific reducing sugar test) was used for other sugars; discontinued as of 2015.
Interferences (false positives): contamination by bleach or oxidizing agents; exposure to air due to improper storage.
Interferences (false negatives or delays): high ascorbic acid; high ketone bodies; refrigerated specimens (enzyme activity decreased); urine must be at room temperature when tested.
Ketone Bodies: Clinical Significance and Testing
Ketone bodies are normal products of fat metabolism but not normally detectable in blood or urine.
Major causes of elevated ketones: diabetes mellitus and starvation; dehydration; fever, vomiting, diarrhea.
In uncontrolled diabetes, urine may contain large amounts of sugar and ketone bodies (diabetic ketoacidosis).
Ketone testing principle: based on Legal (Rothera) test; color reaction with sodium nitroprusside; acetoacetate is the ketone body detected.
Interferences: pigments or drugs that color urine; highly concentrated urine may yield false positives; phthaleins or pH-sensitive substances can cause false positives; improper storage or delayed testing may cause false negatives (conversion of acetoacetate to acetone).
Confirmatory tests: Acetest (sodium nitroprusside-based) for acetone and acetoacetic acid.
Protein: Clinical Significance and Testing
Proteinuria can result from glomerular damage (albuminuria), tubular damage, prerenal or overflow proteinuria (e.g., low-molecular-weight proteins like hemoglobin, myoglobin, or Bence Jones proteins), lower urinary tract disorders, or transient proteinuria (exercise, cold exposure, orthostatic).
Persistent microalbuminuria is associated with diabetes mellitus.
Tamm-Horsfall (uromodulin) protein is produced by renal tubular cells; forms the matrix of urinary casts; casts are often pathologic and associated with proteinuria.
Testing principle (reagent strip): tests for albumin; pH indicators show color changes at specific pH values.
Interferences: false positives from prolonged strip exposure, highly alkaline or buffered urine, or contamination; false negatives when proteins other than albumin predominate.
Historical confirmatory tests: SSA (Sulphosalicylic Acid) precipitation test used to be a standard confirmatory test, but this method has been discontinued due to manual interpretation and availability issues.
Why SSA is discontinued: manual interpretation, low detection rate of true positives, and availability of more sensitive quantitative tests.
Specific Gravity (SG)
SG strips have replaced refractometry in many labs; SG strips measure ionic concentration and do not measure glucose or dyes.
For increased accuracy, when manually testing urine with pH ≤ 6.5, add an adjustment of +0.005 to the SG result.
Note: SG strip results tend to be lower than refractometer results for the same sample; an adjustment step may improve accuracy.
Test principle: measures ionic concentration.
Blood in Urine
Clinical significance: indicates kidney or urinary tract status; tests for RBCs, hemoglobin, and myoglobin via peroxidase activity.
Hematuria: presence of RBCs in urine; may indicate kidney/bladder tumor or stones.
Hemoglobinuria: hemolysis in bloodstream (intravascular).
Myoglobinuria: may result from skeletal muscle injury (e.g., severe trauma).
Differentiation of RBCs, hemoglobin, and myoglobin requires examining the urine sediment under microscopy:
Urine appearance: RBCs present; hemoglobin/myoglobin typically clear and red.
Sediment: RBCs present for hematuria; hemoglobinuria and myoglobinuria usually show few or no RBCs in urine sediment.
Blood testing principle: based on peroxidase-like activity of hemoglobin; false positives with bleach; false negatives with high ascorbic acid (> 25 mg/dL), high specific gravity, or very high nitrite.
Confirmatory testing: microscopic examination of urine sediment.
pH: Significance and Principle
pH describes urine acidity/alkalinity; urine pH ranges from acidic to alkaline depending on diet and metabolism.
Kidney function helps control blood and extracellular fluid pH.
Diet can influence urine pH toward acidity or alkalinity.
pH testing principle: double indicator (methyl red and bromothymol blue) with measurement range 5.0 \, \le\, \text{pH} \, \le\, 9.0.
Interferences: none significant mentioned; fresh urine recommended to prevent false shifts due to bacterial activity.
Fresh urine testing is recommended because bacteria can cause alkaline shifts due to urea conversion to ammonia.
Important practical tip: avoid wetting the strip excessively so that the acid buffer from the protein area runs into the pH area.
Bilirubin: Significance and Testing
Bilirubin is used as an indicator of liver function; normally not detectable in urine because conjugated bilirubin is excreted into bile and converted to urobilinogen in the intestine.
Urine bilirubin testing is performed when abnormal urine color (icteric) is suspected or liver disease is suspected.
Test principle: diazo reaction; strips are bilirubin-specific.
Interferences: bilirubin readings can be difficult to interpret visually; substances in urine can falsely color the sample; false positives from color-causing substances; false negatives from bilirubin oxidation, UV exposure, increased ascorbic acid, or high nitrite; urine should be tested fresh and protected from light.
Confirmatory testing: ICTOTEST tablet (for improved readability)
Urobilinogen: Significance and Testing
Urobilinogen testing is indicated when abnormal urine color or suspected hemolysis or liver disease is suspected.
In the intestine, conjugated bilirubin is converted to urobilinogen by bacteria; some urobilinogen is reabsorbed and filtered by the kidneys.
Normal production: about 1% of bilirubin processed per day; presence in small amounts is common.
Test principle: based on a modified Ehrlich aldehyde reaction.
Interferences:
False positives: highly colored pigments and metabolites; freshly voided urine can yield false positives.
False negatives/decreased results: formalin as a preservative; nitrite > 0.3 mmol/L can reduce readings.
Jaundice: Types and Urine Test Patterns
Prehepatic (hemolytic) jaundice: increased destruction of RBCs leading to increased unconjugated bilirubin and increased urobilinogen.
Hepatic (hepatocellular) jaundice: liver disease; impaired conversion of unconjugated to conjugated bilirubin or impaired transport of conjugated bilirubin into bile.
Posthepatic (obstructive) jaundice: obstruction of the common bile duct; back-up of bilirubin with reduced formation of urobilinogen.
Case-based urine test interpretations (patterns):
Normal: Bilirubin negative; Urobilinogen low/normal.
Prehepatic: Bilirubin negative; Urobilinogen increased.
Hepatic: Bilirubin variable (often increased); Urobilinogen increased or variable.
Posthepatic: Bilirubin increased; Urobilinogen decreased or absent.
Nitrite: Clinical Significance and Testing
Nitrite testing detects certain urinary tract infections (UTIs) caused by Gram-negative bacteria.
Combined testing with leukocyte esterase increases diagnostic value.
Testing principle: Griess test (diazo reaction).
Sample considerations: urine must be retained in the bladder for sufficient time (generally ~4 hours); first-morning specimen is ideal.
Result interpretation: positive or negative; intensity of color does not necessarily reflect infection severity.
Interferences: false positives from medications or in vitro nitrate-to-nitrite conversion due to contamination; false negatives from insufficient time for nitrate-to-nitrite reduction or low dietary nitrate.
Confirmatory tests: microscopic exam, urine culture, Gram stain.
Leukocytes Esterase: Significance and Testing
Indicates leukocyte presence, often associated with UTIs (kidney or bladder) or other infections.
Test principle: detects esterase present in granulocytic leukocytes (primarily neutrophils).
Sensitivity: trace leukocytes can be present in healthy adults; positivity often requires ~5–15 leukocytes per high-power field (HPF).
Interferences: false positives from strong oxidizing agents (chlorine bleach, formalin); preservatives should not be used; false negatives from elevated glucose, high SG, or high albumin.
MLT Verification of Results
MLAs must not report results; verification responsibilities fall to the Medical Laboratory Technician (MLT).
Some automated analyzers connect to LIS and auto-verify results within reference ranges; reported results outside reference ranges or with critical values require MLT verification.
If a result reflexes to a microscopic urinalysis, the MLT prepares the sample for the next testing step (see Unit 5 for more detail).
Urine Disposal and Safety
After collection and testing, dispose of any remaining urine according to local regulations.
Under Bloodborne Pathogens Standard, urine with visible blood is biohazardous and must be handled per standard protocols.
If the specimen is not biohazardous and the device was not used for testing a biohazardous substance, the specimen and device can often be discarded in regular trash after disposal of urine via sink/toilet.
Always check with local regulatory agencies for disposal requirements.
Quick Reference: Reading Time and Result Ranges (Strip Reading Times)
Leukocytes: negative; reading time ~2 minutes.
Nitrite: negative; reading time ~60 seconds.
Protein: range from negative to various positive levels; typical reading times around 60 seconds.
Urobilinogen: ranges and color intensity with reading times ~60 seconds.
pH: reading at ~60 seconds; pH typically ranges 5.0–8.0.
Blood: read at ~60 seconds; color variations indicate negative, trace, small, moderate, large.
Specific Gravity: read at ~45 seconds; typical ranges 1.000–1.030 with increments shown; many strips display 1.000, 1.005, 1.010, 1.015, 1.020, 1.025, 1.030.
Ketone: negative to high values; reading at ~40 seconds.
Bilirubin: negative to varying intensities; reading at ~30 seconds.
Glucose: negative to high; reading at ~30 seconds.
Notes: Reported values include mg/dL or mmol/L for some tests and may be shown as percentage or color intensity; always refer to the specific product’s chart for interpretation.
Case Studies and Interpretive Practice
Case Study 1: Cloudy, yellow, foul-smelling urine with results: Glucose Negative, Ketone Negative, Protein Trace, Specific Gravity 1.020, Blood Negative, pH 7.5, Bilirubin Negative, Urobilinogen Negative, Leukocytes +++, Nitrites +. Presumptive diagnosis: Urinary Tract Infection caused by Gram-negative bacteria.
Case Study 2: Clear, straw-yellow urine with sweet smell and results: Glucose +++, Ketone 8.0 mmol/L, Protein Trace, Specific Gravity 1.015, Blood Negative, pH 6.5, Bilirubin Negative, Urobilinogen Negative, Leukocytes Trace, Nitrites Negative. Presumptive diagnosis: Uncontrolled diabetes mellitus (diabetic ketoacidosis).
Case Study 3: Amber, clear, strong-smelling urine; results: Glucose Negative, Ketone Negative, Protein Trace, Specific Gravity 1.025, Blood Negative, pH 7.5, Bilirubin Negative, Urobilinogen Negative, Leukocytes Negative, Nitrites Negative. Presumptive diagnosis: Likely dehydration or another non-diabetic/low-protein condition; further history needed.
Case Study 4: Red, clear urine; results: Glucose Negative, Ketone Negative, Protein Negative, Specific Gravity 1.020, Blood Negative, pH 7.5, Bilirubin Negative, Urobilinogen Negative, Leukocytes Negative, Nitrites Negative. Presumptive diagnosis: Color change likely due to food or beets; not a clear pathological finding.
Case Study 5: Brown/yellow, clear urine; results: Glucose Negative, Ketone Negative, Protein Negative, Specific Gravity 1.015, Blood Negative, pH 6.0, Bilirubin 35 µmol/L, Urobilinogen Negative, Leukocytes Negative, Nitrites Negative. Presumptive diagnosis: Post-hepatic or obstructive jaundice (elevated bilirubin without bilirubin in urine in this pattern).
Answers to Previous Case Studies (summary):
1) Urinary Tract infection due to Gram-negative bacteria.
2) Uncontrolled diabetes mellitus (diabetic ketoacidosis).
3) Dehydration with need for fluid intake.
4) Possible dietary/color effect (e.g., beets) on urine color.
5) Obstructive (post-hepatic) jaundice.
References and Resources (as listed in the course)
Linné & Ringsrud’s Clinical Laboratory Science: Chapter 13 (pp. 396-412) and related material.
McCall, R.E. Phlebotomy Essentials (7th ed.) Chapter 13.
Mosby’s Canadian Manual of Diagnostic and Laboratory Tests (Chapter 11, pp. 940).
Siemens Multistix 10SG Product Insert and Strip Colour Chart.
Video resources: 67 Seconds on Urinalysis Testing; Urinalysis Dipstick Test.
Additional references: Turgeon, M.L.; MacDonald, S.A.; Pagana K.D. & Pagana T.J. (current ed.).
Note: Some materials may reference additional sources or updated clinical guidelines; always verify against current institutional references.
Quick Reference Formulas and Key Values (Consolidated)
Glucose renal threshold: ext{Renal threshold for glucose} \approx 10\,\mathrm{mmol/L}
Normal SG range: 1.001 \le \text{SG} \le 1.035
Normal urine pH range: 5.0 \le \mathrm{pH} \le 8.0
Urobilinogen reference: 3.2 \le \mathrm{Urobilinogen} \le 17.0\ \mu\mathrm{mol/L}
Bilirubin in urine is typically negative unless indicated otherwise by disease state; bilirubin testing uses a diazo reaction.
Leukocyte esterase threshold: trace to positive depending on leukocyte count; clinically relevant when combined with nitrite results.
Timing reminders: read times per manufacturer instructions; same specimen may yield different results at different times.
Confirmatory tests (where applicable): microscopic urine sediment for blood, ICTOTEST for bilirubin; urine culture for nitrite-positive results with leukocytes.
End of Notes
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