Urine Analysis (Urinalysis) - Pathology Practical

1. Specimen Collection & Evaluation

• Best specimen: First-voided morning urine (most concentrated, best for protein, cells, casts)
• Mid-stream clean catch: Reduces contamination; used for routine analysis and culture
• Volume needed: 10-15 mL for centrifugation
• Timing: Examine within 1-2 hours of collection; refrigerate if delayed (prevents bacterial overgrowth and cell lysis)
• 24-hour urine: Used for quantitative protein, creatinine clearance

2. Physical (Gross) Examination

Color

Clarity / Turbidity

• Clear: Normal
• Turbid / Cloudy: Indicates cells (RBCs, WBCs), casts, crystals, bacteria, mucus, or lipids
• Method: Assessed by reading newsprint through the specimen in a tube

Odor

Volume (24 hours)

• Normal: 1,000-1,500 mL/day
• Polyuria: >2,500 mL/day (diabetes mellitus/insipidus, chronic renal failure)
• Oliguria: <400 mL/day (dehydration, acute renal failure, shock)
• Anuria: <100 mL/day (complete obstruction, severe renal failure)

3. Chemical (Reagent Strip / Dipstick) Examination

pH

• Normal: 4.5-8.0 (average ~6.0, slightly acidic)
• Acid urine: High protein diet, acidosis, fever, diabetes, starvation
• Alkaline urine: Vegetarian diet, UTI (urease-producing organisms), renal tubular acidosis, vomiting

Specific Gravity (SG)

• Normal: 1.003-1.030
• Reflects kidney concentrating ability (also measured by refractometry or urinometer)
• Low SG (<1.010): Dilute urine, diabetes insipidus, renal tubular disease
• High SG (>1.020): Concentrated urine, dehydration, glycosuria, proteinuria
• Fixed SG (1.010): Isosthenuria - indicates severe tubular damage (e.g., chronic glomerulonephritis)

Osmolality

• Normal: 300-900 mOsm/kg
• After dehydration, osmolality should be 3-4x plasma osmolality (~285 mOsm/kg)

Protein

• Normal: Trace or negative (<150 mg/day)
• Dipstick: Primarily detects albumin (false negative for Bence-Jones protein - use SSA test)
• Proteinuria >4 g/day: Nephrotic syndrome
• Mild proteinuria: Glomerulonephritis, renal tubular disease, orthostatic proteinuria
• Bence-Jones proteinuria: Multiple myeloma (heat test - precipitates at 45-60°C, dissolves at 100°C)

Glucose

• Normal: Negative (trace glucose reabsorbed by tubules)
• Glucosuria appears when blood glucose >180 mg/dL (renal threshold)
• Causes: Diabetes mellitus (most common), Cushing's syndrome, renal glycosuria (normal blood glucose, defective tubular reabsorption), pregnancy
• Dipstick: Glucose oxidase method (specific for glucose only)
• Benedict's test / Fehling's test: Detects all reducing sugars (glucose, fructose, galactose, lactose)

Ketones

• Normal: Negative
• Ketonuria: Diabetic ketoacidosis (DKA), starvation, prolonged vomiting, febrile illness, cachexia
• Rothera's test: Ferric chloride or sodium nitroprusside - gives purple-violet color
• Dipstick: Detects acetoacetate and acetone (not beta-hydroxybutyrate)

Blood (Hematuria / Hemoglobinuria)

• Normal: Negative
• Hematuria: >3 RBCs/HPF - from anywhere in urinary tract (kidney, ureter, bladder, urethra)
• Hemoglobinuria: Intravascular hemolysis (malaria, transfusion reactions, PNH)
• Myoglobinuria: Rhabdomyolysis, crush injury
• Note: Dipstick cannot distinguish between RBCs, hemoglobin, and myoglobin - microscopy differentiates

Bilirubin & Urobilinogen

• Bilirubin: Only conjugated (direct) bilirubin appears in urine
• Gmelin test: For bilirubin (green ring at junction)
• Ehrlich's test: For urobilinogen (cherry red color)

Nitrite

• Positive: Indicates gram-negative bacterial infection (bacteria reduce dietary nitrates to nitrites)
• Used along with leukocyte esterase to screen for UTI

Leukocyte Esterase

• Positive: Indicates pyuria (>5 WBCs/HPF), UTI
• False negative: High specific gravity, high glucose, ascorbic acid

4. Microscopic Examination of Urinary Sediment

Procedure

1. Pour 10-15 mL of well-mixed urine into a centrifuge tube
2. Centrifuge at 450g for 5 minutes
3. Discard supernatant (after first performing physical and chemical tests on it)
4. Resuspend sediment in ~0.5 mL remaining supernatant
5. Optionally add supravital stain (Sternheimer-Malbin stain or methylene blue)
6. Load one drop onto a glass slide, apply coverslip
7. Allow to settle for 30-60 seconds
8. Examine under reduced/subdued light (or phase-contrast), varying the fine focus

• Casts: Report per low-power field (LPF) - scan at least 10 LPFs
• RBCs, WBCs, epithelial cells: Report per high-power field (HPF) - average 10 HPFs

A. Red Blood Cells (RBCs)

• Normal: 0-2 RBCs/HPF
• Appear as biconcave discs; in dilute urine may swell (ghost cells); in hypertonic urine may shrink (crenated)
• Dysmorphic RBCs (acanthocytes, "Mickey Mouse ears"): Indicate glomerular origin of bleeding (glomerulonephritis) - caused by mechanical damage passing through the GBM
• Isomorphic RBCs: Non-glomerular bleeding (lower urinary tract - cystitis, calculi, tumors)

B. White Blood Cells (WBCs / Pus Cells)

• Normal: 0-5 WBCs/HPF
• Mostly neutrophils (larger than RBCs, granular cytoplasm, multilobed nucleus)
• Pyuria (>5 WBCs/HPF): UTI, pyelonephritis, glomerulonephritis
• Clumps of WBCs ("pus clumps"): Strongly suggest renal origin (pyelonephritis)
• Glitter cells: WBCs with Brownian movement of granules, seen in hypotonic urine

C. Epithelial Cells

D. Casts

E. Crystals

• Uric acid (yellow-brown rhomboids/rosettes)
• Amorphous urates
• Calcium oxalate (envelope-shaped / "Maltese cross")

• Triple phosphate (coffin-lid shaped) - struvite stones
• Amorphous phosphates
• Calcium carbonate

F. Bacteria, Yeast, Parasites

• Bacteria: If detectable at low power = significant bacteriuria (~100,000 organisms/mL); confirm with culture
• Yeast (Candida): Oval budding cells, may form pseudohyphae; immunocompromised patients, diabetics
• Trichomonas vaginalis: Pear-shaped motile protozoan; STI
• Schistosoma haematobium eggs: Terminal-spined eggs; cause hematuria
• Mucus threads: Long wavy threads; normal in small amounts, not clinically significant

5. Urine Findings in Common Diseases (Summary Table)

Practical Tips (Exam Focus)

1. Always report casts in LPF and cells in HPF
2. RBC casts = glomerulonephritis (most important cast to identify)
3. Oval fat bodies under polarized light = Maltese cross = nephrotic syndrome
4. Broad casts = chronic renal failure (renal failure casts)
5. Cystine crystals (hexagonal) are always pathological
6. Bilirubin in urine = conjugated hyperbilirubinemia (conjugated is water-soluble)
7. Dysmorphic RBCs (acanthocytes) = glomerular disease
8. Dipstick protein test is sensitive for albumin but misses Bence-Jones protein - use SSA test or heat test
9. Fixed SG of 1.010 = isosthenuria = severe tubular damage
10. Supravital stains (Sternheimer-Malbin): Stain WBCs and RTE cells blue-purple, making them easier to identify