Comprehensive Urinalysis Exam 4-6

Benedict's Copper Test: Purpose

Outdated confirmation test of glucose in the urine.

Benedict's Copper Test: Color Progression

Blue to Green to Orange to Red.

Benedict's Copper Test: Principle

Reducing sugars reduce cupric sulfate to cuprous oxide.

Benedict's Copper Test: False Positives

Other reducing sugars (galactose, fructose, and lactose), ascorbic acid, and salicylate medications.

Benedict's Copper Test: False Negatives

Pass-through effect (more analyte than chemical to react with so it goes all the way through positive and then back to a negative color/reduced result)

Clinitest: Purpose

Current glucose confirmation test in the laboratory.

Clinitest: Principle

Reducing sugars reduce cupric sulfate to cuprous oxide using tablets.

Clinitest Tablet Components

Copper sulfate (color change), sodium hydroxide (alkaline environment), citric acid (heat) react with reducing substances in urine to convert cupric acid to cuprous oxide

Clinitest: 5-Drop Method Limit

Detects up to 2% sugar in the urine.

5 drops of urine

If pass through occurs do the 2 drop method

Clinitest: 2-Drop Method Limit

Detects up to 5% sugar in the urine.

Clinitest: False Positives

Other reducing sugars, ascorbic acid (Vitamin C), and salicylate medications.

Clinitest: False Negatives

Pass-through effect.

Clinitest vs. Dipstick Sensitivity

Clinitest is less sensitive than the urine dipstick.

Positive Dipstick / Negative Clinitest

Low glucose levels, expired tablets, or highly pigmented urine causes a false pos on a dipstick

Negative Dipstick / Positive Clinitest

Presence of non-glucose reducing sugars, or expired dipsticks.

Acetest: Purpose

Confirmation of ketones in urine, whole blood, plasma, or serum.

Acetest: Principle

Sodium nitroprusside reacts with acetoacetate in an alkaline environment.

Acetest: Glycine & Lactose Role

Glycine detects acetone; lactose causes the color change.

Ketone Not Detected by Acetest

Beta-hydroxybutyrate.

Acetest: False Positives

Medications with sulfhydryl groups or highly pigmented urine.

Acetest: False Negatives

Bacterial degradation, expired/improperly stored tablets, or pass-through effect.

Differentiating Sulfa Drugs on Acetest

Add glacial acetic acid; purple fades if due to sulfa drugs.

Ictotest: Purpose

Confirmation test of bilirubin in the urine.

Ictotest: Principle

Azocoupling reaction with diazonium salt using absorbent mats.

Ictotest Tablet Components

Diazonium salt, sulfosalicylic acid, and sodium bicarbonate.

Ictotest vs. Dipstick Sensitivity

Ictotest is more sensitive than the urine dipstick.

Ictotest: False Positives

Highly pigmented urine or chlorpromazine medications.

Ictotest: False Negatives

High ascorbic acid/nitrites, improper storage, or cold urine.

Sulfosalicylic Acid (SSA) Test: Purpose

Confirmation of protein in the urine.

SSA Test: Principle

Performed on clear urine obtained from centrifugation

Mixing centrifuged urine supernatant with 3% SSA causes proteins to

precipitate.

Turbidity is assessed for results

SSA Test: Protein Sensitivity

Picks up any protein but is most sensitive to albumin.

SSA Test: False Positives

High concentration of antibiotics or X-ray contrast media.

SSA Test: False Negatives

Dilute urine or highly buffered alkaline urine.

SSA Test: Urine Turbidity Interference

Initial turbidity can cause false positives or false negatives if misread.

Microscopic Prep: Recommended Urine Volume

12 mL

Microscopic Prep: Volume < 12 mL Protocol

Use 6 mL and double the final results.

Microscopic Prep: Volume < 6 mL Protocol

Use 3 mL, bypass centrifugation, and test unspun sample.

Microscopic Prep: Centrifugation Parameters

1500 rpm for 5 minutes.

Microscopic Prep: Resuspension Volume

1 mL of sediment is resuspended after decanting supernatant.

Sediment Microscopy: Objective for Cells

Observed and counted on low power (10x).

Sediment Microscopy: Objective for Casts

Observed, identified, and counted on high power.

Sediment Microscopy: Objective for Crystals

Counted on low power; identified on high power.

Sediment Microscopy: Number of Fields to View

10 to 15 microscopic fields.

Brightfield Microscopy: Urinalysis Characteristics

Most common but hardest to see low-contrast sediment; requires constant focusing up and down

Phase-Contrast Microscopy: Urinalysis Role

Preferred method for translucent, low-refractive sediment, but expensive.

Polarizing Microscopy: Urinalysis Role

Used to identify highly refractile elements by illuminating them with polarizing light.

Interference-Contrast Microscopy: Urinalysis Role

Provides a 3D image of sediment; too expensive for routine urinalysis.

Sternheimer-Malbin Stain: Alternate Name & Use

Kova Stain; most common stain for cellular structures and formed elements.

Crystal violet and safranin

Toluidine Blue Stain (0.5%): Purpose

Enhances nuclear detail of cells in sediment.

Acetic Acid in Urine Sediment: Purpose

Lyses red blood cells and enhances white blood cell nuclear patterns.

Fat Stains (Oil Red O / Sudan III): Purpose

Confirms the presence of neutral fats in urine.

Gram Stain: Purpose

Stain used when there is lots of bacteria present in the urine to see if gram positive or negative

Prussian Blue Stain: Purpose

Confirms the presence of hemosiderin (high iron storage).

Hansel Stain: Purpose & Components

Methylene blue and Eosin-Y in methanol; used to identify eosinophils.

Patients with allergic reactions

Urine RBCs: Normal Range & Objective

Less than 3 per high power field (hpf); viewed on high power.

Hematuria: Definition

The presence of red blood cells in the urine.

Ghost Cell: Definition

Swollen, colorless RBC in hypotonic urine that has lost its hemoglobin.

RBC Appearance: Hypertonic vs. Hypotonic Urine

Hypertonic: Shrunk and crenated.

Hypotonic: Swollen, colorless, or lysed.

Pathological causes of RBCs in the Urine

Hemolytic anemia, Hemophilia, Transfusion reactions

Non-pathological causes of RBCs in the Urine

Strenuous exercise, menses, medications (blood thinners)

Positive Blood Dipstick

RBCs are lysed, hemoglobin released, or interference with reagent strip

Negative Blood Dipstick

RBC lookalikes, interference with reagent strip

Differentiating RBCs from Yeast/WBCs

Acetic acid lyses RBCs but not WBCs or yeast; RBCs are refractile.

Urine WBCs: Normal Range & Objective

Less than 8 per high power field (hpf); viewed on high power.

Pyuria: Definition

The presence of white blood cells in the urine.

What is the most common type of WBC found in urine?

Neutrophils

Glitter Cell: Definition

WBC showing light play on granules; seen in hypotonic/dilute urine.

Pathological causes of WBCs in urine

Inflammation, Chlamydia, Gonorrhea, Yeast, Trichomonas

Non-pathological causes of WBCs in urine

Strenuous exercise and fever

Positive Leukocyte Esterase

WBCs are lysed

Negative Leukocyte Esterase

WBC lookalikes (renal tubular cells, large RBC), or lymphocytes are the WBC present or not enough for the dipstick to pick up

Squamous Epithelial Cells: Origin

Lining of vaginal canal and female urethra; distal male urethra.

Transitional (Caudate) Epithelial Cells: Origin

Lining of renal pelvis, bladder, and upper urethra.

Renal Tubular Epithelial (RTE) Cells: Origin & Significance

Distal tubule/collecting ducts; >2/hpf indicates renal necrosis.

Characteristics of Squamous Epithelial Cells

Abundant and irregular cytoplasm, central nucleus about the same size of an RBCs

1:9-10 ratio

Flagstone cells

Low Power objective

Characteristics of Transitional Epithelial Cells

Small than squamous, round, polyhedral or caudate

Central nucleus

1:3-5 ratio

High power objective

Pathogenic in large numbers

Upper UTI, Urinary procedures due to catheter

Characteristics of Renal Tubular Epithelial Cells

Round or slightly elongated

Slightly larger than a WBC

Single round nucleus that is off center

1:2 ratio

Pathological: >2/hpf is indicative of renal necrosis which can be caused by pyelonephritis, toxin reaction, viral infection, kidney transplant rejection, glomerulonephritis

Primary Cast Protein: Name & Alias

Uromodulin, also known as Tamm-Horsfall protein.

Four Factors of Cast Formation

Decreased urine flow

Increased acidity

Increased solute concentration

Increased plasma proteins

General Characteristics of Urinary Casts

Formed primarily within the distal convoluted tubule and collecting duct and excreted in the urine

Presence of casts will be accompanied by proteinuria

Shapes are usually cylindrical with parallel sides and rounded ends

Hyaline Casts: Characteristics & Significance

Most common; uromodulin-only; refractive index close to urine

Difficult to see with a brightfield microscope

Pathological cause of a Hyaline cast

Renal disease and heart disease

Non-Pathological cause of Hyaline cast

Strenuous exercise, dehydration, heat exposure, and emotional stress

Granular Casts: Composition

Contains granules throughout the matrix of the cast: the larger the granules the worse off the patient

Easily viewed on a brightfield

Consists of uromodulin, metabolized renal tubular cells, and disintegrated cellular casts

Pathological cause of Granular casts

Renal disease

Non-Pathological cause of Granular casts

Stress and Strenuous exercise

Oval Fat Bodies: Definition & Appearance

Renal tubular cells that have absorbed fat; highly refractile and circular.

Contains free fat globules, oval bodies, or both in a hyaline or granular matrix

Fats in the urine is always pathogenic

RBC Cast Composition and Appearance

Refractile and range in color: yellow, brown, and reddish brown

Contains uromodulin and RBCs embedded as either whole, crenated or lysed

As RBC casts age they become hemoglobin casts

Conditions that cause RBC Casts

Glomerulonephritis and occasionally strenuous exercise

Composition and Appearance of WBC casts

Refractile with granules and multi-lobed nuclei

Contains uromodulin and WBCs

WBC Casts: Clinical Significance

Indicates infection/inflammation within the nephron; highly associated with pyelonephritis.

Composition and Appearance of Renal epithelial casts

Contains uromodulin and renal tubule epi cells within the cast

Difficult to differentiate from WBC casts unless contrast microscopy is used

Conditions that cause Renal epithelial casts

Indicated tubular injury

May be associated with heavy metal and chemical or drug induced toxicity, viral infection, allograft rejection, and kidney infection

Composition and Appearance of Waxy Casts

Refractile and homogeneously smooth

Ends may appear sharp, blunt, uneven and even cracked

Waxy Casts: Clinical Significance

Represents extreme stasis (>48 hours); signifies chronic renal failure.

What are the 3 factors that contribute to form urinary crystals

Changes in pH or temp

Concentration of urine solutes

Slow flow of urine through tubules

Normal Acid Crystals (pH < 7)

Amorphous urates, uric acid, acid urates, monosodium urates, calcium oxalate, hippuric acid.

Amorphous Urates: Macroscopic Appearance

Yellow-brown granules; forms a pink "brick dust" precipitate due to uroerythrin.

Often present in shapeless clumps

Uric Acid Crystals: Appearance & pH

Diamond, cube, or rosette shapes

Yellow/orange-brown

Only present if pH < 5.7 otherwise present in its salt form (acid urates and monosodium urates)

Acid Urates

Appears as small balls or spheres

Usually yellow brown in color

Uric acid salt