URINALYSISBy

Roderick D. Balce, RMT

ANATOMY OF THE KIDNEY

Afferent arteriolesEfferent arteriolesPeritubular capillariesProximal convoluted tubuleDistal convoluted tubuleVasa rectaAscending loop of HenleDescending loop of Henle

RENAL FUNCTIONS

1. Renal Blood Flow

2. Glomerular Filtration Glomerulus GFR = 120 ml/min Factors Affecting Glomerular Filtration

a. Cellular structure Capillary Wall Membrane Basement Membrane Visceral Epithelium

b. Hydrostatic pressure Blood Colloidal Oncotic Pressure Capsular Pressure

c. Renin-Angiotensin-Aldosterone Mechanism

3. Tubular Reabsorption Mechanisms involved:

a. Active Transport Actively transported: Glucose,

Amino acids Salts

Chloride Sodium

Renal threshold –important in distinguishing between: Excess solute filtration Renal tubular damage

b. Passive TransportPassively Transported: Water

Urea Sodium

c. Counter-Current MechanismOsmotic Gradient of Medulla

Vasopressin

4. Tubular SecretionMajor Functions:

a. Elimination of waste products not filtered by the glomerulus

b. Regulation of acid-base balance through secretion of H ions

Renal Function Tests1. Glomerular Filtration Test

Clearance testa. Ureab. Radioisotopec. B2 microglobulind. Creatinine C = UV/P Normal values: Creatinine Clearance: F = 75-112 ml/m

M = 85-125 ml/m Plasma Creatinine = 0.5-1.5 mg/dle. Inulin

2. Tubular Reabsorption Tests a. Fishberg Testb. Mosenthal Test

c. Osmometry

3. Tubular Secretion and Renal Blood Flow Tests

a. PAHb. PSPc. Indigo Carmine Test

Renal DiseasesA. Glomerular Disorders:1. Glomerulonephritis – blood, protein, and casts

in urinea. Acute Poststreptococcal – caused by

deposition of immune complexes and antibodies in the glomerular

membrane following group A streptococcal infection.

b. Rapidly Progressive ( Crescentic) – more serious, can lead to renal failure; arises as a result of another form of glomerulonephritis or an immune systemic disorder.

c. Good Pasture’s Syndrome – attachment of cytotoxic autoantibody (anti-glomerular

basement membrane antibody) to the glomerular and alveolar basement membranes during viral respiratory infections.

Lab Findings:proteinuria, hematuria, RBC casts

2. Vasculitis - immune-mediated disoders affecting the systemic vascular system resulting to

glomerular damage.

a. Wegener’s granulomatosis – granuloma- producing inflammation of the small blood vessels in the lungs and kidney.The antibody causing the damage is the anti-neutrophilc cytoplasmic antibody (ANCA) Lab findings: hematuria, proteinuria, rbc casts,

and increased BUN and crea

b. Henoch-Schonlein Purpura – characterized by a decrease in platelets that causes disruption of vascular integrity Lab findings: heavy proteinuria and hematuria

with rbc casts

Wegener’s Granulomatosis

3. Immunoglobulin A nephropathy

- deposition of immune complexes on the glomerular membrane resulting from increased levels of serum IgA Lab findings: macroscopic/microscopic

hematuria and increased IgA

4. Membranous Glomerulonephritis

- thickening of the glomerular membrane due to deposition of IgG immune complexes. Lab findings: microscopic hematuria and

increased IgG and protein excretion

5. Membranoproliferative Glomerulonephritis – immune mediated disorder characterized by cellular proliferation in capillary walls or glomerular basement membrane.

Lab findings: hematuria, proteinuria, and decreased serum complement levels

6. Chronic Glomerulonephritis – glomerular damage as a result of other renal disorders leads to marked decrease in renal functions and eventually to renal failure Lab findings: hematuria, proteinuria, glucosuria,

varieties of casts including broad cast, markedly decreased GFR with increased BUN and crea levels, and electrolyte imbalance

7. Nephrotic Syndrome – disruption in the electrical charges in the basal lamina and podocytes, producing a less tightly connected barrier. Lab Findings: massive proteinuria, low albumin, high

serum levels of lipids, and pronounced edema

8. Minimal Change Disease – podocytes appear to be less tightly fitting allowing increased filtration of protein; seen in children

following allergic reaction and immunization. Lab Findings: edema, heavy proteinuria, transient

hematuria

9. Focal Segmental Glomerulosclerosis- only a certain number and areas of

glomeruli are affected. The disease is caused by disruption of the podocytes associated with analgesics and heroin abuse and AIDS. Immunoglobulins M and C3 are seen in undamaged

glomeruli.•Lab Findings: heavy proteinuria, microscopic hematuria

II. Tubulointerstitial Disorders 1. Acute Tubular Necrosis – damage to the

renal tubular cells by toxic agents or ischemia

2. Fanconi’s Syndrome – generalized failure of tubular reabsorption in the PCT;

may be hereditary or acquired

3. Cystitis – ascending bacterial infection of the bladder

4. Acute Pyelonephritis – infection of the upper urinary tract involving the interstitium

and tubules due to interference of urine flow to the bladder, reflux of urine from the bladder, or untreated cystitis

5. Chronic Pyelonephritis – recurrent infection of the tubules and interstitium caused

by structural abnormalities affecting urine flow

6. Acute Interstitial Nephritis – inflammation of the renal interstitium associated with allergic reaction to medications

C. Vascular Disorders1. Renal Failure – may be gradual progression

from the original disorder to chronic renal failure or end-stage renal disease.

2. Renal Lithiasis – deposition of renal calculi or kidney stones in the calyces and pelvis of the kidney, ureters and urinary bladder.

Chemical Composition of Renal Calculi:

a. Calcium oxalate or phosphate

b. Magnesium ammonium phosphate

c. Uric acid

d. Cystine

Lab Findings: microscopic hematuria

Composition of Urine

1. water

2. analytes organic inorganic

3. hormones, vitamins, medications, formed elements, etc.

Timed Specimen: 24-hour Quantitative chemical tests,

hormone studies

12-hour Addis count

2-hr Postprandial Diabetic monitoring

Afternoon Specimen Urobilinogen determination

Glucose Tolerance Test Accompaniment to blood samples in GTT

Types of Urine Specimens

Random Routine screeningFirst Morning Routine screening

Pregnancy TestsOrthostatic Proteinuria

Fasting/Second Morning Diabetic screening / monitoring

Catheterized Bacterial cultureMidstream clean-catch Routine screening

Bacterial cultureSuprapubic aspiration Bladder urine for bacterial

cultureCytology

Three-glass collection Prostatic infectionDrug Specimen Drug testing

Voiding cystourethrogramExamining bladder function by injecting dye that is

visible on X-rays through a catheter to fill the bladder. X-rays are taken while the bladder is full and while the patient is urinating to determine if

fluid is forced out of the bladder through the urethra (normal) or up through the ureters into the

kidney (vesicoureteral reflux).

DRUG SPECIMEN COLLECTION

Chain of Custody (COC) Form the process that provides the

documentation of proper sample identification from the time of collection to the receipt of laboratory results

a standardized form that documents that the specimen collected by the patient is the same one that is analyzed and reported

Urine Specimen Collection

- the most vulnerable part of a drug testing program

- may be witnessed or unwitnessed; a same-gender collector is required in witnessed collection

Required Volume - 30-45 ml taken within 4 minutes

Temperature - 32.5ºC to 37.7ºC

Procedure: The collector washes hands, wears

gloves, adds bluing agent or dye to the toilet water reservoir, and tapes the toilet lid and faucet handles.

The donor provides identification from employer.

The collector completes step 1 COC Form and has the donor sign the form.

The donor leaves all his belongings outside, washes hands, and receives a specimen cup.

The collector remains in the restroom, outside the stall (unwitnessed) listening for unauthorized water use.

The donor hands specimen cup to the collector and the latter checks the urine for abnormal color and for the required amount.

The collector checks the temperature using a temperature strip, records the reading on the COC Form (step 2). If the temperature is out of range, recollection is needed.

With the donor watching, the collector seals the capped bottle with identification strips (COC step 3) covering both sides of the cup. The seals contain the date and time.

The donor initials the seals and completes COC step 4 after which, the collector accomplishes COC step 5.

Each time the specimen is handled, transferred or stored, every individual must be identified, the date and purpose of the change must be recorded, and specific instructions on labeling, packaging or transport must be followed.

Methods of Preservation1. Physical (Refrigeration)

2. Chemical Phenol – causes an odor change Toluene –not effective for bacteria and molds Thymol crystals – preserves glucose and

sediments well Formalin – excellent sediment preservative Boric acid – preserves protein well Sodium fluoride – good for drug analysis Sacomanno’s fixative – preserves cellular

elements

Changes in Unpreserved Urine Color - modified

or darkened

Increased:OdorpHNitriteBacteria

Decreased:ClarityGlucoseKetonesBilirubinUrobilinogenRed blood cellsWhite blood cellsCasts

Clinical Utilities of Routine Urinalysis

Indicators of the State of the Kidney or Urinary TractAppearance Specific GravityChemical testsLeukocyte EsteraseUrinary Sediment

Indicators of Metabolic and Other Conditions or DiseasepHAppearanceGlucose and KetonesBilirubinUrobilinogen

Indications of Other Systemic (Nonrenal) Conditions or DiseaseHemoglobinMyoglobinLight-chain proteinsPorphobilinogen

PHYSICAL EXAMINATION

I. VolumeAverage daily output: 1,200-1,500 mlVariations:

a. Polyuria Indications: Diabetes mellitus

Diabetes insipidus

b. Diuresisc. Oliguriad. Anuriae. Nocturia

II. Specific Gravity Performed using:

a. Urinometer Corrections done

b. Refractometer Instrument Calibration

c. Reagent strip

d. Harmonic Oscillation Densitometry

Variations in S.G.:a. Hypersthenuriab. Hyposthenuriac. Isosthenuria

III. pH

IV. Color• Pigments

Variations in Color: Colorless Pale yellow Dark yellow Orange Red/

Reddish brown Amber/Orange

Yellow green/

Yellow brown Green Blue green Milky white Pink/Red Brown/Black

V. Odor Normal: Aromatic

Variations in Urine Odor:

Amoniacal/Putrid/Foul

Mercaptan Fecaloid Fruity, sweet Maple syrup Sulfur odor

Mousy Rancid Sweaty feet Rotting fish Cabbage Bleach

VI. TransparencyUrine ClarityClear No visible particulates, transparent

Hazy Few particulates, print easily seen through urine

Cloudy Many particulates, print blurred through urine

Turbid Print cannot be seen through urine

Milky May precipitate or be clotted

Nubecula

Nonpathologic Causes of Urine Turbidity

Epithelial cells Normal crystals Bacteria (old urine) Semen, prostatic fluid Fecal contamination Radiographic contrast media, mucus,

talcum powder Vaginal creams

Pathologic Causes of Urine Turbidity

Red blood cells White blood cells Bacteria Yeasts Nonsquamous epithelial cells Abnormal crystals Casts Lymph fluid/Chyle Lipids Fecal matter

CHEMICAL ANALYSIS

I. pH Normal values: average-6; random-4.5-8.0;

fasting-5.5-6.5 Clinical Significance

a. respiratory or metabolic acidosis/ketosisb. respiratory or metabolic alkalosisc. renal tubular acidosisd. renal calculi formatione. treatment of UTIf. precipitation/identification of crystalsg. determination of unsatisfactory specimen

Causes of Acid and Alkaline Urine

Acid Urine Alkaline Urine Emphysema Hyperventilation Diabetes mellitus Vomiting Starvation Renal tubular acidosis Dehydration Presence of urease-

producing bacteria Diarrhea Vegetarian diet Presence of Old specimens

acid-producing

bacteria High protein diet Cranberry juice Medications

II. Protein Most indicative of renal disease Normal urine contains <10 mg/dl or 100 mg/24

hours Types of Proteinuria (according to the amount of

protein excreted per day)

Degree of Grams Excreted Proteinuria per 24 hours

Mild <1 g/dayModerate 1- 4 g/day Heavy >4 g/day

Types of Proteinuria (according to cause)

1. Pre-renal Intravascular hemolysisMuscle injurySevere infection and inflammationMultiple myeloma

2. Renal a. Glomerular

Conditions that cause damage to the glomerulus Orthostatic or Postural Proteinuria

b. Tubular

• Fanconi’s Syndrome

• Toxic agents/Heavy metals

• Severe viral infections

3. Post-renala. lower UTI/inflammation

b. injury/trauma

c. menstrual contamination

d. prostatic fluid/spermatozoa

e. vaginal secretions

Tests for Albumin Heat and Acetic Acid Test

Positive Results: 1+ - diffused cloud

2+ - granular cloud

3+ - distinct flocculi

4+ - large flocculi Reagent Strips SSA/Cold Protein Precipitation

Correlation of Reagent Strip and SSA Results

(+) Rgt. Strip, (-) SSA = albumin present

(+) Rgt. Strip, (+)SSA = proteinuria

(-) Rgt. Strip, (+) SSA = BJP, globulins, etc.

III. Glucose Normal concentration: 15 mg/dl Melituria Glycusoria Clinical significance

a. DMb. endocrine disordersc. pancreatitis, carcinoma, cystic fibrosis,

hemochromatosisd. CNS disorderse. disturbance in metabolismf. liver diseaseg. renal glycusoriah. drugs

Glycusoria without hyperglycemia

a. renal tubular dysfunction

b. tubular necrosis

c. Fanconi’s syndrome

Tests for Glucose

1.Benedict’s Test – general test for glucose and other reducing sugars

2. Clinitest – subject to interference from other reducing sugars

• Pass-though phenomenon

3. Glucose Oxidase – specific for glucose

Correlation of Glucose Oxidase and Clinitest

GOD Clinitest Interpretation 1+ neg small amount of glucose

present

4+ neg possible oxidizing agent interference on rgt

strip

neg 1+ non-glucose reducing substance

present

IV. Ketones Represents 3 intermediate products of fat

metabolism:

a. acetone

b. acetoacetic acid

c. β-hydroxybutyric acid

Accumulation in blood leads to:

a. electrolyte imbalance

b. dehydration

c. acidosis

d. diabetic coma

Clinical significance:a. DM

b. insulin dose monitoring

c. diabetic acidosis

d. starvation/fasting

e. weight reduction/dieting/strenuous exercise

f. vomiting

g. malabsorption/pancreatic disorders

h. inborn error of amino acid metabolism

V. BloodHematuria

Clinical Significance:a. renal calculib. glomerulonephritisc. tumorsd. traumae. pyelonephritisf. toxic chemicalsg. strenuous sxerciseh. menstrual contamination

Hemoglobinuria

Clinical significance:a. hemolytic anemia

b. transfusion reaction

c. PNH

d. severe burns and infections

e. malaria

f. strenuous exercise

Myoglobinuria

Clinical significance:a. muscular trauma/crush syndrome

b. prolonged coma

c. convulsions

d. muscle-wasting diseases

e. alcoholism/overdose

f. drug abuse

g. extensive exertion

Tests to Differentiate Hemoglobin and Myoglobin

Ammonium Sulfate Method Hb is precipitated by ammonium sulfate

Absorption Spectrophotometry Immunodiffusion Technique Protein Electrophoresis

Differentiation of Hematuria, Hemoglobinuria, and Myoglobinuria

Finding Red Cells Hemoglobin Myoglobin

Rgt. Strip Positive Positive Positive

Microscopic (RBCs)

Present Absent or few

Absent or few

Urine Appearance Cloudy red Clear red Clear red

Plasma Appearance

Normal Pink to red Normal

Total serum CK Normal Slight elevation

Marked elevation

Total serum LD Normal Elevated Elevated

LD1 and LD2 Normal Elevated Normal

LD4 and LD5 Normal Normal Elevated

VI. Bilirubin Clinical Significance

Hepatitis, cirrhosis, other liver disorders Biliary obstruction

Tests Foam-Shake Test Oxidation Test – acidic oxidation of bilirubin

into rainbow array of colors

Diazotization Test Ictotest Reagent Strip

VII. Urobilinogen found in urine in small amounts

Clinical Significance Early detection of liver disease Hemolytic disorders Hepatitis, cirrhosis, carcinoma

Tests Ehrlich’s Tube Test Schwartz-Watson Differentiation Test Reagent Strip

Urobilinogen Ehrlich-reactive Substances

Porphobilinogen

Chloroform Extraction

Urine (Top layer)

Chloroform(Bottom layer)

Colorless

Red

Red

Colorless

Red

Colorless

Butanol Extraction

Butanol (Top layer)

Urine (Bottom layer)

Red

Colorless

Red

Colorless

Colorless

Red

Watson-Schwartz Test Interpretation

Laboratory Findings in Various Types of Jaundice

Blood Urine Urine

Bilirubin Bilirubin Urobilinogen

Normal 0–1.3 mg/dL Negative ≤1 mg/dL

Hemolytic Increased Negative +++

Hepatic Increased + or - ++

Obstructive Normal +++ Normal

Urobilin – brown pigment which result from oxidation of urobilinogen upon exposure to light; tested when

urobilinogen is negative Porphyrins – cyclic compounds derived from ALA

Clinical Significance: Porphyrias Tests for Porphyrinuria

Ehrlich Test – requires the addition of acetylacetone prior to performing the test

Fluorescence – involves extraction into a mixture of glacial HAc and ethyl acetate and

examination of solvent layer

faint blue fluorescence = (-) violet, pink, or red = (+)

VIII. Nitrite Clinical Significance

Cystitis Pyelonephritis Evaluation of antibiotic therapy Monitoring of patients at high risk for

UTI Screening of urine culture specimens

IX. Leukocyte EsteraseClinical Significance

Bacterial and nonbacterial UTI Inflammation of the urinary tractScreening of urine culture

specimens

Urine Reference ValuesProperty Reference Value

Color Yellow

Transparency Clear

pH 5-7

Specific Gravity 1.001-1.035

Protein (albumin) Negative or Trace

Blood (hemoglobin) Negative

Nitrite Negative

LE Negative

Glucose Negative

Ketones Negative

Bilirubin Negative

Urobilinogen ≤1 mg/dL

Confirmatory Urinalysis TestsSubstance Test

Protein SSA Test

Blood Microscopic Examination

Hemoglobin, Myoglobin Centrifugation; then test supernatant with rgt.

strip for blood

Hemosiderin Rous Test

Glucose Clinitest

Bilirubin Ictotest

Urobilinogen Watson-Schwartz Test

Porphobilinogen Hoesch Test

Ascorbic Acid EM Quant

SPECIAL URINALYSIS SCREENING TESTS

Amino Acid Disorders

1. Phenylalanine-Tyrosine Disorders

a. Phenylketonuria

Tests: Ferric Chloride Test Phenistix DPNH Robert Guthrie Bacterial Inhibition

Test

b. Tyrosyluria Tests:

Millon’s Test Nitrosonaphthol Test Ferric Chloride Test Phenistix DPNH

c. Melanuria Tests:

Sodium nitroprusside Test - interference due to a red color from

acetone and creatinine can be avoided by adding glacial HAc

Ferric Chloride Test Acetest Blackberg and Wanger Test Ehrlich’s Test

d. Alkaptonuria

Tests: Ferric chloride Test Alkali Test Addition of silver nitrate or ammonium

hydroxide Benedict’s Test or Clinitest

2. Branched Chain Amino Acid Disorders

a. Maple Syrup Urine Disease Tests:

DNPH Amino Acid Chromatography Ferric Chloride Test Nitrosonaphthol Acetest

b. Organic Acidemias Isovaleric Acidemia

DNPH Acetest Chromatography

Propionic Acidemia DNPH Acetest

Methylmalonic Acidemia DNPH Acetest p-nitroaniline

3. Tryptophan Disorders

a. Indicanuria

Clinical Significance: Obstruction Presence of bacteria Malabsorption syndrome

b. Hartnup’s Disease production of blue color when indican is

oxidized upon exposure to air

c. 5-HIAA Tests:

Ferric Chloride Test Sjoerdsma Test.

Clinical Significance: Elevated in malignancy involving

Argentaffin cells

4. Cystine Disordersa. Cystinuria Tests:

+ Cystine crystals Cyanide nitroprusside

b. CystinosisTests: Cyanide nitroprusside Clinitest

c. HomocystinuriaTests: Cyanide nitroprusside Silver nitroprusside

Mucopolysaccharide Disorders Hurler’s Syndrome Hunter’s Syndrome San Filippo’s Syndrome

Tests: Acid Albumin CTAB Turbidity Test Metachromatic Staining Spot Test

Purine DisordersLesch-Nyhan Disease

lack of the enzyme HGPT Manifestations: severe motor defects,

mental retardation, tendency toward self-destruction, gout and renal calculi, orange sand in diapers

MICROSCOPIC EXAMINATION

Types of Microscope Bright Field – objects appear dark against a light

background Phase Contrast – works by retardation of light

rays diffused by the object in focus

Polarizing – used to confirm the identification of fat droplets, oval fat bodies, and fatty casts

Interference Contrast – object appears bright against a dark background but without the diffraction halo associated with Phase Contrast Microscope

Microscopic Sediment StainsSternheimer-Malbin0.5% Toluidine BlueSudan IIIOil Red OPrussian BlueHansel Stain

Reference Values for Urine Sediment

Constituent Reference Value

Red Blood Cells 0-2/hpf

White Blood Cells 0-5/hpf (female>male)

Casts 0-2 hyaline/hpf

Squamous Epithelial Cells Few/hpf

Transitional Epithelial Cells Few/hpf

Renal Tubular Epithelial Cells Few/hpf

Bacteria Negative

Yeast Negative

Abnormal crystals Negative

Standardization of Procedure

1. Urine Volume – 12 mL

2. Time of Centrifugation – 5 minutes

3. Speed of Centrifugation Relative Centrifugal Force of 400 g

4. Volume of Sediment Examined – 20 μL

5. Reporting Format

Reporting System for Urine SedimentAverage Number per Low-Power Field

Casts Neg 0-2 2-5 5-10 10-25 25-50 >50

Abnormal crystals Neg 0-2 2-5 5-10 10-25 25-50 >50

Squamous ECs Few Moderate Many

Mucus Present

Average Number per High-Power Field

Red Blood Cells 0-2 2-5 5-10 10-25 25-50 50-100 >100

White Blood Cells 0-2 2-5 5-10 10-25 25-50 50-100 >100

Normal Crystals Few Moderate Many

Epithelial Cells Few Moderate Many

Miscellaneous Few Moderate Many

Sperm Present

Organized Sediments

Red Blood Cells

•Normal appearance

•N.V.

•Variations in shape and appearance

•Clinical Significance

Non-glomerular hematuria: RBCs are uniform in size and shape but

show two populations of cells because a small number have lost

their hemoglobin pigment.

Glomerular hematuria: RBCs are small and vary in size, shape, and

hemoglobin content.

White Blood Cells

Amoeboid

Bacteria

•Normal appearance

•N.V.

•Clinical Significance

•Glitter cells

•Eosinophils

•Mononuclear cells

Squamous Epithelial Cells

• Clue Cells

Transitional Epithelial Cells

smaller than squamous cells, spherical, caudate, or polyhedral with central nucleus

Clinical Significance Transitional that appear singly, in pairs,

or in clumps Transitional cells with abnormal

morphology

Renal Tubular Epithelial Cells

•Clinical Significance

•Oval Fat Bodies

•Bubble Cells

Casts

Mechanisms of Cast Formation

Hyaline Cast

•Strenuous exercise, dehydration, heat exposure, emotional stress

•Acute glomerulonephritis, pyelonephritis, chronic renal disease, CHF

Red Blood Cell Cast

WBC Cast

•Epithelial Cell Cast

Coarsely Granular Cast

Finely Granular Cast

Waxy Cast

•Fatty Cast

•Broad Cast

Bacteria

Yeast Cells

There are no PMNs seen suggestive of contamination and

not of UTI.

Mucus Thread

Unorganized Sediments

CrystalsNormal Crystals in Acidic Urine

1. Amorphous Urates2. Uric Acid3. Sodium Urates4. Calcium Sulfates5. Calcium Oxalate6. Hippuric Acid

Amorphous Urates

Uric Acid

Dihydrate Calcium Oxalate

Normal Crystals in Alkaline Urine

1. Amorphous Phosphate

2. Calcium Carbonate

3. Ammonium biurate

4. Calcium Phosphate

5. Triple Phosphate

Amorphous Phosphate

Calcium Carbonate

Ammonium biurate

Calcium phosphate

Triple phosphate

Abnormal Crystals of Metabolic Origin

Cystine Cholesterol Leucine Tyrosine Bilirubin Hemosiderin

Abnormal Crystals of Iatrogenic Origin

Sulfonamides Radiographic

contrast media Ampicillin Acyclovir Indinavir sulfate

Cystine

Cholesterol

Leucine

Tyrosine

Sulfa Crystals

Starch