Transcript Urinalysis
Urinalysis
Course: IDPT 5005 School of Medicine, UCDHSC
Francisco G. La Rosa, MD [email protected]
Associate Professor, Department of Pathology University of Colorado Denver Health Sciences Programs, Denver, Colorado 80045
Specimen Collection
– – – – –
First morning voiding (most concentrated) Record collection time Type of specimen (e.g. “clean catch”) Analyzed within 2 hours of collection Free of debris or vaginal secretions
Clean Catch
Specimen Collection
Supra-pubic Needle Aspiration
Types of Analysis
− − − − −
Macroscopic Examination Chemical Analysis (Urine Dipstick) Microscopic Examination Culture (not covered in this lecture) Cytological Examination
Macroscopic Examination
Odor:
− − − − −
Ammonia-like: Foul, offensive: Sweet: Fruity: Maple syrup-like: Color:
− − − − − −
Colorless Deep Yellow Yellow-Green Red Brownish-red Brownish-black (Urea-splitting bacteria) Old specimen, pus or inflammation Glucose Ketones Maple Syrup Urine Disease Diluted urine Concentrated Urine, Riboflavin Bilirubin / Biliverdin Blood / Hemoglobin Acidified Blood (Actute GN) Homogentisic acid (Melanin)
Macroscopic Examination
Turbidity:
− − − −
Typically cells or crystals.
Cellular elements and bacteria will clear by centrifugation.
Crystals dissolved by a variety of methods (acid or base).
Microscopic examination will determine which is present.
Chemical Analysis
Chemical Analysis
Urine Dipstick Glucose Bilirubin Ketones Specific Gravity Blood pH Protein Urobilinogen Nitrite Leukocyte Esterase
The Urine Dipstick:
Glucose Negative Trace (100 mg/dL) + (250 mg/dL) ++ (500 mg/dL) +++ (1000 mg/dL) ++++ (2000+ mg/dL) Chemical Principle
Glucose Oxidase
Glucose + 2 H 2 O + O 2 ---> Gluconic Acid + 2 H 2 O 2
Horseradish Peroxidase
3 H 2 O 2 + KI ---> KIO 3 + 3 H 2 O Read at 30 seconds RR: Negative
Uses and Limitations of Urine Glucose Detection Significance
– –
Diabetes mellitus.
Renal glycosuria.
Limitations
– – –
Interference: reducing agents, ketones.
Only measures glucose and not other sugars.
Renal threshold must be passed in order for glucose to spill into the urine.
Other Tests
–
CuSO 4 test for reducing sugars.
Detection of Reducing Sugars* by CuSO 4 Sugar - Galactose - Fructose Intolerance, etc.
- Lactose - Pentoses - Maltose Disease
(s)
Galactosemias Fructosuria, Fructose Lactase Deficiency Essential Pentosuria Non-pathogenic * NOT Sucrose because it is not a reducing sugar
Urine versus Blood Glucose ++ + trace Negative 200 400 600 800 Blood Glucose (mg/dL) 1000
Microscopic Examination
General Aspects Preservation - Cells and casts begin to disintegrate in 1 - 3 hrs. at room temp.
- Refrigeration for up to 48 hours (little loss of cells).
Specimen concentration - Ten to twenty-fold concentration by centrifugation.
Types of microscopy - Phase contrast microscopy - Polarized microscopy - Bright field microscopy with special staining (e.g., Sternheimer-Malbin stain)
Microscopic Examination
Abnormal Findings Per High Power Field (HPF) (400x)
– – – –
> 3 erythrocytes > 5 leukocytes > 2 renal tubular cells > 10 bacteria Per Low Power Field (LPF) (200x)
– – –
> 3 hyaline casts or > 1 granular cast > 10 squamous cells (indicative of contaminated specimen) Any other cast (RBCs, WBCs) Presence of:
– – –
Fungal hyphae or yeast, parasite, viral inclusions Pathological crystals (cystine, leucine, tyrosine) Large number of uric acid or calcium oxalate crystals
Microscopic Examination
Cells Erythrocytes “Dysmorphic” vs. “normal” (> 10 per HPF) Leukocytes - Neutrophils (glitter cells) - Eosinophils More than 1 per 3 HPF Hansel test (special stain) Epithelial Cells - Squamous cells - Renal tubular epithelial cells - Transitional epithelial cells Indicate level of contamination Few are normal Few are normal - Oval fat bodies Abnormal, indicate Nephrosis
Microscopic Examination
RBCs
Microscopic Examination
RBCs
Microscopic Examination
WBCs
Microscopic Examination
Squamous Cells
Microscopic Examination
Tubular Epithelial Cells
Microscopic Examination
Transitional Cells
Microscopic Examination
Transitional Cells
Microscopic Examination
Oval Fat Body
Microscopic Examination
LE Cell
Microscopic Examination
Bacteria & Yeasts Bacteria - Bacteriuria Yeasts - Candidiasis Viruses - CMV inclusions More than 10 per HPF Most likely a contaminant but should correlate with clinical picture.
Probable viral cystitis.
Microscopic Examination
Bacteria
Microscopic Examination
Yeasts
Microscopic Examination
Yeasts
Microscopic Examination
Cytomegalovirus
Microscopic Examination
Casts Erythrocyte Casts: Leukocyte Casts: Degenerating Casts: - Granular casts - Hyaline casts - Waxy casts - Fatty casts (oval fat body casts) Glomerular diseases Pyuria, glomerular disease Nonspecific (Tamm-Horsfall protein) Nonspecific (Tamm-Horsfall protein) Nonspecific Nephrotic syndrome
Microscopic Examination
Casts
Microscopic Examination
RBCs Cast - Histology
Microscopic Examination
RBCs Cast
Microscopic Examination
RBCs Cast - Histology
Microscopic Examination
WBCs Cast
Microscopic Examination
Tubular Epith. Cast
Microscopic Examination
Tubular Epith. Cast
Microscopic Examination
Granular Cast
Microscopic Examination
Hyaline Cast
Microscopic Examination
Waxy Cast
Microscopic Examination
Fatty Cast
Significance of Cellular Casts
Erythrocyte Casts Leukocyte Casts Bacterial Casts Single Erythrocytes Single Leukocytes Single Bacteria Verrier-Jones & Asscher, 1991.
Microscopic Examination
Crystals - Urate Ammonium biurate Uric acid - Triple Phosphate - Calcium Oxalate Amino Acids Cystine Leucine Tyrosine - Sulfonamide
Microscopic Examination
Calcium Oxalate Crystals
Microscopic Examination
Calcium Oxalate Crystals Dumbbell Shape
Microscopic Examination
Triple Phosphate Crystals
Microscopic Examination
Urate Crystals
Microscopic Examination
Leucine Crystals
Microscopic Examination
Cystine Crystals
Microscopic Examination
Ammonium Biurate Crystals
Microscopic Examination
Cholesterol Crystals
Cytological Examination
Staining:
–
Papanicolau
–
Wright’s
–
Immunoperoxidase
–
Immunofluorescence
Cytology: Normal
Cytology: Normal
Cytology: Reactive
Cytology: Reactive
Cytology: Polyoma
(Decoy Cell)
Cytology: Polyoma
Immunoperoxidase to SV40 ag (Decoy Cell)
Cytology: TCC Low Grade
Cytology: TCC Low Grade
Cytology: TCC High Grade
Cytology: TCC High Grade
Cytology: Squamous Cell Ca.
Cytology: Renal Cell Ca.
Cytology: Prostatic Carcinoma
Questions ?
Disclaimer: The images and texts presented in this slide show are solely for educational purposes and not intended for commercial or pecuniary benefit. The images have been obtained from Dr. La Rosa’s personal collection, from text books used during the teaching of this chapter, and from published articles and educational works. Reproduction of these images can be done only for educational use.
Reference: USA Copyright Law, Section 110, “Limitations on exclusive rights: Exemption of certain performances and displays”).
[ Download ] the USA Copyright Law version, October 2007.
Chemical Analysis
Urine Dipstick Glucose Bilirubin Ketones Specific Gravity Blood pH Protein Urobilinogen Nitrite Leukocyte Esterase
The Urine Dipstick:
Glucose Negative Trace (100 mg/dL) + (250 mg/dL) ++ (500 mg/dL) +++ (1000 mg/dL) ++++ (2000+ mg/dL) Chemical Principle
Glucose Oxidase
Glucose + 2 H 2 O + O 2 ---> Gluconic Acid + 2 H 2 O 2
Horseradish Peroxidase
3 H 2 O 2 + KI ---> KIO 3 + 3 H 2 O Read at 30 seconds RR: Negative
Uses and Limitations of Urine Glucose Detection Significance
– –
Diabetes mellitus.
Renal glycosuria.
Limitations
– – –
Interference: reducing agents, ketones.
Only measures glucose and not other sugars.
Renal threshold must be passed in order for glucose to spill into the urine.
Other Tests
–
CuSO 4 test for reducing sugars.
Detection of Reducing Sugars* by CuSO 4 Sugar - Galactose - Fructose Intolerance, etc.
- Lactose - Pentoses - Maltose Disease
(s)
Galactosemias Fructosuria, Fructose Lactase Deficiency Essential Pentosuria Non-pathogenic * NOT Sucrose because it is not a reducing sugar
Urine versus Blood Glucose ++ + trace Negative 200 400 600 800 Blood Glucose (mg/dL) 1000
The Urine Dipstick:
Bilirrubin Negative + (weak) ++ (moderate) +++ (strong) Chemical Principle Bilirubin + Diazo salt
--------->
Azobilirubin Read at 30 seconds RR: Negative
Uses and Limitations of Urine Bilirrubin Detection Significance - Increased direct bilirubin (correlates with urobilinogen and serum bilirubin) Limitations - Interference: prolonged exposure of sample to light - Only measures direct bilirubin--will not pick up indirect bilirubin Other Tests - Ictotest (more sensitive tablet version of same assay) - Serum test for total and direct bilirubin is more informative
The Urine Dipstick:
Ketones Negative Trace (5 mg/dL) + (15 mg/dL) ++ (40 mg/dL) +++ (80 mg/dL) ++++ (160+ mg/dL) Chemical Principle Acetoacetic Acid + Nitroprusside ------> Colored Complex Read at 40 seconds RR: Negative
Uses and Limitations of Urine Ketone Detection Significance - Diabetic ketoacidosis - Prolonged fasting Limitations - Interference: expired reagents (degradation with exposure to moisture in air) - Only measures acetoacetate not other ketone bodies (such as in rebound ketosis).
Other Tests - Ketostix (more sensitive tablet version of same assay) - Serum glucose measurement to confirm DKA
The Urine Dipstick:
Specific Gravity 1.000
1.005
1.010
1.015
1.020
1.025
1.030
Chemical Principle X + + Polymethyl vinyl ether / maleic anhydride ---------------> X + -Polymethyl vinyl ether / maleic anhydride + H + H + interacts with a Bromthymol Blue indicator to form a colored complex.
Read up to 2 minutes RR: 1.003-1.035
Uses and Limitations of Urine Specific Gravity Significance - Diabetes insipidus Limitations - Interference: alkaline urine - Does not measure non-ionized solutes (e.g. glucose) Other Tests - Refractometry - Hydrometer - Osmolality measurement (typically used with water deprivation test)
The Urine Dipstick:
Blood Negative Chemical Principle Trace (non-hemolyzed) Lysing agent to lyse red blood cells Moderate (non-hemolyzed) Diisopropylbenzene dihydroperoxide + Tetramethylbenzidine Trace (hemolyzed) Colored Complex + (weak) ++ (moderate) Read at 60 seconds RR: Negative Analytic Sensitivity: 10 RBCs +++ (strong)
Uses and Limitations of Urine Blood Detection Significance - Hematuria (nephritis, trauma, etc) - Hemoglobinuria (hemolysis, etc) - Myoglobinuria (rhabdomyolysis, etc) Limitations - Interference: reducing agents, microbial peroxidases - Cannot distinguish between the above disease processes Other Tests - Urine microscopic examination - Urine cytology
The Urine Dipstick:
pH 5.0
6.0
6.5
7.0
7.5
8.0
8.5
Chemical Principle H + interacts with: Methyl Red (at high concentration; low pH) and Bromthymol Blue (at low concentration; high pH), to form a colored complexes (dual indicator system) Read up to 2 minutes R.R.: 4.5-8.0
Uses and Limitations of Urine pH Detection Significance - Acidic (less than 4.5): metabolic acidosis, high-protein diet - Alkaline (greater than 8.0): renal tubular acidosis (>5.5) Limitations - Interference: bacterial overgrowth (alkaline or acidic), “run over effect” effect of protein pad on pH indicator pad Other Tests - Titrable acidity - Blood gases to determine acid-base status
pH Run Over Effect Glucose Bilirubin Ketones Specific Gravity Blood pH Protein Urobilinogen Nitrite Leukocyte Esterase Buffers from the protein area of the strip (pH 3.0) spill over to the pH area of the strip and make the pH of the sample appear more acidic than it really is.
The Urine Dipstick:
Protein Negative Chemical Principle
“Protein Error of Indicators Method”
H H H Pr Pr Pr Trace + (30 mg/dL) ++ (100 mg/dL) +++ (300 mg/dL) ++++ (2000 mg/dL) H H H Tetrabromphenol Blue (buffered to pH 3.0) Pr Pr Pr Pr Pr Pr Pr Read at 60 seconds RR: Negative Pr Pr H + H H + H + + H H + +
Causes of Proteinuria Functional - Severe muscular exertion - Pregnancy - Orthostatic proteinuria Renal - Glomerulonephritis - Nephrotic syndrome - Renal tumor or infection Pre-Renal - Fever - Renal hypoxia - Hypertension Post-Renal - Cystitis - Urethritis or prostatitis - Contamination with vaginal secretions
Nephrotic Syndrome (> 3.5 g/dL in 24 h) Primary - Lipoid nephrosis (severe) - Membranous glomerulonephritis - Membranoproliferative glomerulonephritis Secondary - Diabetes mellitus (Kimmelsteil-Wilson lesions) - Systemic lupus erythematosus - Amyloidosis and other infiltrative diseases - Renal vein thrombosis
Uses and Limitations of Urine Protein Detection Significance - Proteinuria and the nephrotic syndrome.
Limitations - Interference: highly alkaline urine.
- Much more sensitive to albumin than other proteins (e.g., immunoglobulin light chains).
Other Tests - Sulfosalicylic acid (SSA) turbidity test.
- Urine protein electrophoresis (UPEP) - Bence Jones protein
Proteins in “Normal” Urine Protein Albumin Tamm-Horsfall Immunoglobulins Secretory IgA Other TOTAL % of Total 40% 40% 12% 3% 5% 100% Daily Maximum 60 mg 60 mg 24 mg 6 mg 10 mg 150 mg
The Urine Dipstick:
Urobilinogen 0.2 mg/dL 1 mg/dL 2 mg/dL 4 mg/dL 8 mg/dL Chemical Principle Urobilinogen + Diethylaminobenzaldehyde (Ehrlich’s Reagent) -------> Colored Complex Read at 60 seconds RR: 0.02-1.0 mg/dL
Uses and Limitations of Urobilinogen Detection Significance - High: increased hepatic processing of bilirubin - Low: bile obstruction Limitations - Interference: prolonged exposure of specimen to oxygen (urobilinogen ---> urobilin) - Cannot detect low levels of urobilinogen Other Tests - Serum total and direct bilirubin
The Urine Dipstick:
Nitrite Chemical Principle Negative Positive Nitrite + p-arsenilic acid -------> Diazo compound Diazo compound + Tetrahydrobenzoquinolinol ----------> Colored Complex Read at 60 seconds RR: Negative
Uses and Limitations of Nitrite Detection Significance - Gram negative bacteriuria Limitations - Interference: bacterial overgrowth - Only able to detect bacteria that reduce nitrate to nitrite Other Tests - Correlate with leukocyte esterase and - Urine microscopic examination (bacteria) - Urine culture
The Urine Dipstick:
Leukocyte Esterase Negative Trace + (weak) ++ (moderate) +++ (strong) Chemical Principle Derivatized pyrrole amino acid ester 3-hydroxy-5-phenyl pyrrole 3-hydroxy-5-phenyl pyrrole + diazo salt -------------> Colored Complex Read at 2 minutes RR: Negative Analytic Sensitivity: 3-5 WBCs
Uses and Limitations of Leukocyte Esterase Detection Significance - Pyuria - Acute inflammation - Renal calculus Limitations - Interference: oxidizing agents, menstrual contamination Other Tests - Urine microscopic examination (WBCs and bacteria) - Urine culture
Urinalysis Disease Diagnosis
Case 1
Diluted urine, request a voided urine in the morning If persisting low SG, possible diabetes insipida A microscopic may give negative results Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
Negative Negative Negative 1.001
Negative 5.5
Negative 0.2 mg/dL Negative Negative A 35-year old man undergoing routine pre employment drug screening.
Physical characteristics: Clear.
Microscopic: Not performed.
Drugs Identified: None.
Questions: - What is your differential diagnosis?
- What would you do next to confirm your suspicion?
- Would you order a microscopic analysis on this sample?
Case 2
Possible gallbladder or hepatic disease.
No hemolytic anemia. Perform bilirubins in serum Microscopic unlikely to provide additional info Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
Negative +++ A 42 year old woman presents with “dark urine” Negative 1.020
Physical characteristics: Red-brown.
Microscopic: Not performed.
Negative 5.5
Questions: Negative 0.2 mg/dL - What is your differential diagnosis?
- Could this be a case of hemolytic anemia?
- How would you rule it out?
- What tests would you order next? Why?
- Would you order a microscopic analysis?
Negative Negative
Case 3
Possible UTI, request culture and antibiotic sensitivity Negative Nitrite test: Gram positive bacteria Lower SG may show less number of cells and bacteria Un-common diagnosis in this type of patient Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
Negative Negative Negative 1.030
+++ 6.5
Trace 1.0 mg/dL Negative +++ A 42-year old man presents painful urination Physical characteristics: dark red, turbid Microscopic: leukocytes = 30 per HPF RBCs = >100 per HPF Bacteria = >100 per HPF Questions: - What is your suspected diagnosis?
- What would you do next?
- What do you make of the nitrite test?
- How would the microscopic exam differ if the S.G. were 1.003?
- Is this a common diagnosis for this type of patient?
Case 4
Diabetes May be decompensated and with ketoacidosis Ketones should become negative after treatment Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
++ Negative Trace A 27-year old woman presents with severe abdominal pain.
1.015
Negative 6.0
Negative 1.0 mg/dL Physical characteristics: clear-yellow.
Microscopic: Not performed.
Questions: - What is the most likely diagnosis?
- What do you make of the ketone result?
- What do you expect to happen to the ketone measurement when treatment begins?
Negative Negative
Case 5
Glomerulonephritis RBC casts reveals renal cortex involvement RBC cast are not always present in GN Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
Negative Negative Negative 1.015
+++ 6.5
+ 1.0 mg/dL Negative Negative 8-year old boy presents with discolored urine Physical characteristics: Red, turbid.
Microscopic: erythrocytes = >100 per HPF (almost all dysmorphic) Red cell casts present.
Questions: - What is the most likely diagnosis in this case?
- Does the presence of red cell casts help you in any way?
- If the erythrocytes were not dysmorphic would that change your diagnosis?
Case 6
“Functional” proteinuria?
Microscopic may reveal a few leukocytes Request protein concentration in 24 h urine Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
Negative Negative Negative 1.010
Negative 5.0
+ 0.2 mg/dL Negative Negative 22-year old man presenting for a routine physical required for admission to medical school Physical characteristics: Yellow Microscopic: Not performed Questions: - What is your differential diagnosis?
- Would you order a microscopic analysis on this sample?
- What would you do next to confirm the diagnosis?
Common Findings in: Acute Tubular Necrosis Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
Decreased + / + / Microscopic:
• •
Renal tubular epithelial cells Pathological casts
Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
Increased Increased Common Findings in: Acute Glomerulonephritis Microscopic:
• • •
Erythrocytes (dysmorphic) Erythrocyte casts Mixed cellular casts
Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
Decreased Increased Increased Common Findings in: Chronic Glomerulonephritis Microscopic:
•
Pathological casts (broad waxy casts, RBCs)
Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
Trace Positive Positive Common Findings in: Acute Pyelonephritis Microscopic:
• • • •
Bacteria Leukocytes Leukocyte, granular, and waxy casts Renal tubular epithelial cell casts
Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
++++ Common Findings in: Nephrotic Syndrome Microscopic:
• • •
Oval fat bodies Fatty casts Waxy casts
Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
+ Common Findings in: Eosinophilic Cystitis Microscopic:
• •
Numerous eosinophils (Hansel’s stain) NO significant casts.
Glucose Bilirubin Ketones S.G.
Blood pH Protein Urobilinogen Nitrite L.E.
+ Common Findings in: Urothelial Carcinoma Microscopic:
•
Malignant cells on urine cytology (urine sample should be submitted separately to cytology, void or 24 hrs.)
Acknowledgment: Dr. Brad Brimhall