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Microscopic_Examination_of_Urine_pdf (1).pptx
- 2. Preparation oftheUrine Sediment Freshly voided urine midstream clean-catch specimen 10 and 15 mL (12ml) Centrifuged for 5 mins @ 400RCF 0.5-1.0 mL left after decantation Represents urinary sediments Agitate Place a drop (20 μL or 0.02 mL) of centrifuged urine in a slide + glass cover slip (22 X22 mm ) Read microscopically
- 3. Examination oftheUrine Sediment Observe minimum of 10 fields [both low & high PF] LPF (10x) detect casts and to ascertain the general composition of the sediment HPF (40x) Identification casts have a tendency to locate near the edges of the cover slip (LPFscanning around the cover slip perimeter)
- 4. Examination oftheUrine Sediment Bright field microscopy (unstained sediments) Artifacts are larger than urinary sediments Reportingof Microscopic Sediments Sediment Reporting Comment Casts Lpf 10 fields RBCs, WBCs Hpf Epithelial cells, crystals, and other elements Semi- quantitative terms rare, few, moderate, and many, or as 1+, 2+, 3+, and 4+, following laboratory format as to lpf or hpf use
- 5. Reportingof Microscopic Sediments Manner of Comment Reporting Occasional 0-1, 0-3/HPFat 10 fields Few 0-4, 2-5/HPFat 10 fields Some 3-5, 8-10/HPFat 10fields Moderate More than 10/HPFat 10 fields Many Abundant/HPF TNTC Too numerous to count (loaded) Examination of theUrine Sediment
- 6. Sediment ExaminationTechniques (Stains) Stain Action Function Sternheimer- Malbin (Crystal violet &Safranin O) Delineates structure and contrasting colors of the nucleus and cytoplasm Identifies WBCs, epithelial cells, and casts Toluidine blue Enhances nuclear detail Differentiates WBCs and renal tubular epithelial (RTE) cells 2 % acetic acid Lyses RBCs and enhances nuclei of WBCs Distinguishes RBCs from WBCs, yeast, oil droplets, and crystals
- 7. Sediment ExaminationTechniques (Stains) Stain Action Function Lipid Stains:Oil Red O and Sudan I I I Stains triglycerides and neutral fats orange-red Identifies free fat droplets and lipid-containing cells and casts Gram stain Differentiates gram- positive and gram- negative bacteria Identifies bacterial casts Hansel stain Methylene blue and eosin Y stains eosinophilic granules Identifies urinary eosinophils Prussianblue stain Stains structures containing iron Identifies yellow-brown granules of hemosiderin in cells and casts
- 8. Expected StainingReactions of Sediment Constituents Elementsin UrinarySediment UsualDistinguishingColor of Stained Elements Comments RBCs Neutral—pink to purple Acid—pink (unstained) Alkaline—purple Hyaline casts Pale pink or pale purple Very uniform color; slightly darker than mucous threads Coarse granular inclusion casts Dark purple granules in purple matrix Finely granular inclusion casts Fine dark purple granules in pale pink or pale purple matrix
- 9. Expected Staining Reactions of Sediment Constituents Elementsin Urinary Sediment UsualDistinguishingColor of Stained Elements Comments Nuclei Cytoplasm WBCs(dark-staining cells) Purple Purple granules Glittercells (Sternheimer- Malbin positive cells) Colorless or light blue Pale blue or gray Some glitter cells exhibit brownian movement Renal tubular epithelial cells Dark shade of blue-purple Light shade of blue-purple Bladder tubular epithelial cells Blue-purple Light purple Squamous epithelial cells Dark shade of orange-purple Light purple or blue
- 10. Expected Staining Reactions of Sediment Constituents Elementsin Urinary Sediment UsualDistinguishing Color ofStained Elements Comments Waxy casts Pale pink or pale purple Darker than hyaline casts, but of a pale even color; distinct broken ends Fatinclusion casts Fat globules unstained in a pink matrix Rare; presence isconfirmed if examination under polarized light indicates double refraction Red cell inclusion casts Pink to orange-red Intact cells can be seen in matrix Blood (hemoglobin) casts Orange-red No intact cells
- 11. Expected StainingReactions of Sediment Constituents Elementsin Urinary Sediment UsualDistinguishing Color ofStained Elements Comments Bacteria Motile: do not stain Nonmotile: stain purple Motile organisms are not impaired Trichomon as vaginalis Light blue-green Motility isunimpaired in fresh specimens when recommended volumes of stain are used; immobile organisms also identifiable Mucus Pale pink or pale blue Background Pale pink or pale purple
- 12. Cytodiagnostic Urine Testing preparation of permanent slides using cytocentrifugation Staining with Papanicolaou stain Detection of malignancies of the lower urinary tract First morning specimen provides more definitive information about renal tubular changes associated with transplant rejection; viral, fungal, and parasitic infections; cellular inclusions; pathologic casts; and inflammatory conditions
- 13. UrinalysisMicroscopic Techniques Technique Function Bright-field microscopy Used for routine urinalysis Phase-contrast microscopy Enhances visualization of elements with low refractive indices, such as hyaline casts, mixed cellular casts, mucous threads, and Trichomonas Polarizing microscopy Aids in identification of cholesterol in oval fat bodies, fatty casts, and crystals
- 14. UrinalysisMicroscopic Techniques Technique Function Dark-field microscopy Aids in identification of Treponema pallidum Fluorescence microscopy Allows visualization of naturally fluorescent microorganisms or those stained by a fluorescent dye Interference- contrast Produces a three-dimensional microscopy-image and layer- by-layer imaging of a specimen
- 15. Sediment Constituents Red Blood Cell (RBC) Pointof reference Description Appearance • Non-nucleated biconcave disks • Crenated in hypertonic urine • Ghost cells in hypotonic urine • Dysmorphicwith glomerular membrane damage Sources of Identification error • Yeast cells • Oil droplets • Air bubbles Reporting Average number per 10 hpfs Complete urinalysis correlation • Color • Reagent strip blood reaction
- 18. Sediment Constituents WhiteBlood Cell (WBC) Pointof reference Description Appearance • Larger than RBCs (12μdiameter) • Granulated, multilobed neutrophils • Glitter cells in hypotonic urine • Mononuclear cells with abundant cytoplasm Sources of Identification error • Renal tubular epithelial cells Reporting Average number per 10 hpfs Complete urinalysis correlation • Leukocyte esterase • Nitrite • Specific gravity • pH
- 21. Sediment Constituents Epithelial Cells (Squamous) Pointof reference Description Appearance • Largest cells in the sediment with abundant, irregular cytoplasm and prominent nucleii Sources of Identification error • Rarely encountered, folded cells may resemble casts Reporting • Rare, few, moderate, or many per lpf Complete urinalysis correlation • Clarity
- 24. Sediment Constituents Epithelial Cells (Transitional/Urothelial) Pointof reference Description Appearance • Spherical, polyhedral, or caudate with centrally located nucleus Sources of Identification error • Spherical forms resemble RTEcells Reporting • Rare, few, moderate, or many per hpf Complete urinalysis correlation • Clarity; blood, ifmalignancy- associated
- 25. Transitional (Urothelial) Epithelial Cells
- 26. Sediment Constituents Epithelial Cells (Renal Tubular) Pointof reference Description Appearance • Rectangular, columnar, round, oval or, cuboidal with an eccentric nucleus possibly bilirubin-stained or hemosiderin-laden Sources of Identification error • Spherical transitional cells Granular casts Reporting • Average number per 10 hpfs Complete urinalysis correlation • Leukocyte esterase and nitrite(pyelonephritis) • Color *Clarity • Protein *Bilirubin (hepatitis) • Blood
- 27. Renal Tubular Epithelial Cells
- 28. Sediment Constituents Oval Fat Bodies Pointof reference Description Appearance • Highly refractile RTEcells Sources of Identification error • Confirm with fat stains and polarized microscopy Reporting • Average number per hpf Complete urinalysis correlation • Clarity • Blood • Protein • Free fat droplets/fatty casts
- 29. Oval FatBodies
- 30. Sediment Constituents Bacteria Pointof reference Description Appearance • Small spherical and rod-shaped structures Sources of Identification error • Amorphous phosphates and urates Reporting • Few, moderate, or many per hpf, the presence of WBCs may be required Complete urinalysis correlation • pH • Nitrite • LE • WBCs
- 31. Bacteria
- 32. Sediment Constituents Yeast Pointof reference Description Appearance • Small, oval, refractile structures with buds and/or mycelia Sources of Identification error • RBCs Reporting • Rare, few, moderate, or many per hpf, the presence of WBCs may be required Complete urinalysis correlation • Glucose • LE • WBCs
- 33. Yeast
- 34. Sediment Constituents Parasites (Trichomonas) Pointof reference Description Appearance • Pear-shaped, motile, flagellated Sources of Identification error • WBCs • renal tubular epithelial cells Reporting • Rare, few, moderate, or many per hpf Complete urinalysis correlation • LE • WBCs
- 35. Parasites
- 36. Sediment Constituents Spermatozoa Pointof reference Description Appearance • Tapered oval head with long, thin tail Sources of Identification error • None Reporting • Present, based on laboratory protocol Complete urinalysis correlation • Protein
- 37. Spermatozoa
- 38. Sediment Constituents Mucus Pointof reference Description Appearance • Single or clumped threads with a low refractive index Sources of Identification error • Hyaline casts Reporting • Rare, few, moderate, or many per lpf Complete urinalysis correlation • None
- 39. Mucus
- 40. UrineCasts Unique to kidney Formed within the lumen of DCT & CD Provides microscopic view of condition w/in nephron Lpf detection/Scanning around cover slip edge Low refractive index observe in subdued light Reporting: Ave. #/10 lpf
- 41. Casts (Composition and Formation) Tamm-Horsfall protein - major constituent of cast - stress & exercise Albumin, immunoglobulin Protein gel urine-flow stasis, acidity, Na+, Ca++ Tapered end formed at the junction of ALH& DCT -cylindroids Cylindruria presence of urinary cast
- 42. Hyaline Casts Pointof reference Description Appearance • Colorless homogenous matrix Sources of Identification error • Mucus, fibers, hair, increased lighting Reporting • Average number per lpf Complete urinalysis correlation • Protein -Blood (exercise) • Color (exercise) Clinical significance • Glomerulonephritis • Pyelonephritis • Chronic renal disease • Congestive heart failure • Stress and exercise
- 43. Hyaline Casts
- 44. RBCCasts Pointof reference Description Appearance • Orange-red color, cast matrix containing RBCs Sources of Identification error • RBC clumps Reporting • Average number per lpf Complete urinalysis correlation • RBCs • Blood • Protein Clinical significance • Glomerulonephritis • Strenuous exercise
- 45. RBCCasts
- 46. WBCCasts Pointof reference Description Appearance • Cast matrix containing WBCs Sources of Identification error • WBC clumps Reporting • Average number per lpf Complete urinalysis correlation • WBCs Protein LE Clinical significance • Pyelonephritis • Acute interstitial nephritis
- 47. WBCCasts
- 48. Bacterial Casts Pointof reference Description Appearance • Bacilli bound to protein matrix Sources of Identification error • Granular casts Reporting • Average number per lpf Complete urinalysis correlation • WBC cast -WBCs • LE -Nitrite • Protein -Bacteria Clinical significance • Pyelonephritis
- 49. EpithelialCell Casts Pointof reference Description Appearance • RTEcells attached to protein matrix Sources of Identification error • WBC cast Reporting • Average number per lpf Complete urinalysis correlation • Protein RTEcells Clinical significance • Renal tubular damage
- 51. FattyCasts Pointof reference Description Appearance Fat droplets and oval fat bodies attached to protein matrix Sources of Identification error Fecal debris Reporting • Average number per lpf Complete urinalysis correlation • Protein • Free fat droplets • Oval fat bodies Clinical significance • Nephrotic syndrome • Toxic tubular necrosis • Diabetes mellitus -Crush injuries
- 52. Fatty Casts
- 53. Granular Casts Pointof reference Description Appearance Coarse and fine granules in a cast matrix Sources of Identification error • Clumps of small crystals • Columnar RTEcells Reporting • Average number per lpf Complete urinalysis correlation • Protein • Cellular casts • RBCs & WBCs Clinical significance • Glomerulonephritis • Pyelonephritis • Stress and exercise
- 54. Granular Casts
- 55. WaxyCasts Pointof reference Description Appearance • Highly refractile cast with jagged ends and notches Sources of Identification error • Fibers and fecal material Reporting • Average number per lpf Complete urinalysis correlation • Protein • Cellular casts • Granular casts • WBCs & RBCs Clinical significance • Stasis of urine flow • Chronic renal failure
- 56. WaxyCasts
- 57. Broad Casts Pointof reference Description Appearance • Wider than normal cast matrix Sources of Identification error • Fecal material, fibers Reporting • Average number per lpf Complete urinalysis correlation • Protein • WBCs & RBCs • Granular casts • Waxy casts Clinical significance • Extreme urine stasis • Renal failure
- 58. UrineCrystals detect the presence of the relatively few abnormal types -liver disease -inborn errors of metabolism -renal damage Reporting: rare, few, moderate, or many per hpf Abnormal crystals average number per lpf
- 59. UrineCrystals formed by the precipitation of urine solutes Subject to: changes in temperature, solute concentration, and pH (affect solubility) Rapidly precipitates at low temperature Presence of crystals in fresh urine high sp. Gr. pH determine type of crystal present *Organic & iatrogenic compounds ppts. Inacidic pH *inorganic salts ppts in neutral & alkaline sol’n *except: CaOx ppts in acidic & neutral urine
- 60. Normal Crystal Seen inAcidic Urine Crystal Color &Shape Solubility Appearnce Uric Acid Yellow-brown (rhombic/4- sided/rosette) Alkali Soluble Amorphous urates Brick dust or yellow brown granules Alkali and heat Calcium oxalate [Acid/neutral (alkaline)] Colorless (envelopes, oval, dumbbell, octahedral) Dilute HCl
- 61. Uricacid crystal
- 62. Amorphous urates
- 63. Calcium oxalate
- 64. Normal Crystal Seen inNeutral/Alkaline Urine Crystal Color &Shape Solubility Appearnce Amorphous phosphates White–colorless (granular/white ppt) Dilute acetic acid Calcium phosphate Colorless (flat rectangular plates/thin prisms often in rosette formations) Dilute acetic acid
- 67. Normal Crystal Seen inAlkaline Urine Crystal Color &Shape Solubility Appearnce Triple phosphate Colorless (prism shape “coffin lids”) Dilute acetic acid Ammonium biurate Yellow-brown (spicule-covered spheres “thorny apples”) Acetic acid with heat Calcium carbonate Colorless (dumbbells/sphe rical) Gas from acetic acid
- 69. Ammonium biurate
- 71. Abnormal Crystals seen inAcid Urine Crystal/Colo r/shape Comment Solubility Appearnce Cystine • Colorless • Hexagon al plates • Cystinuria metabolic disorder of renal tubules • Confirmation test cyanide- nitroprusside test Ammonia, dilute HCl Cholesterol • Colorles s notched plates • Seen in refrigerated specimen (droplet form lipids) • Nephrotic syndrome Chloroform
- 72. Crystal/Color Comments Solubility Appearan c e Bilirubin • Yellow • clumped needles or granules • present in hepatic disorders • positive chemical test result for bilirubin would be expected Acetic acid, HCl, NaOH, ether, chloroform Radiograph ic dye • Colorless • Cholestero l crystal • Markedly high specific gravity when measured by refractometer 10% NaOH Abnormal Crystals seen inAcid Urine
- 73. Cystine
- 74. Cholesterol
- 75. Bilirubin
- 76. Radiographic dye
- 77. Abnormal Crystalsseen inAcid/Neutral Urine Crystal/Color Comments Solubility Appearance Leucine • Yellow-brown • Spheres (concentric circles and radial striations) • Crystals Associated WithLiverDisorders • Presence should be accompanied by tyrosine crystals Hot alkali or alcohol Tyrosine • Colorless–yellow • Fine needle (clump or rosette form) • seen in conjunction with leucine crystals in specimens with positive chemical test results for bilirubin • May be encountered in inherited disorders of amino-acid metabolism Alkali or heat
- 78. Abnormal Crystals seen inAcid/Neutral Urine Crystal/Color Comments Solubility Appearance Sulfonamides • Varied (olorless to yellow-brown) • Needles, rhombics, whetstones, sheaves of wheat, and rosettes • Seen in inadequate patient hydration • possibility of tubular damage ifcrystals are forming in the nephron Acetone Ampicillin • Colorless • needles (form bundles following refrigeration • Seen in massive doses of this penicillin compound without adequate hydration Refrigera- tion forms bundles
- 79. Leucine
- 80. Tyrosine
- 81. Sulfonamide
- 82. Ampicillin
- 83. UrinarySediment Artifacts Contaminants from improper collection starch, oil droplets, air bubbles, pollen grains, fibers, and fecal contamination often very highly refractile or occur in a different microscopic plane Not necessarily to be reported
- 84. UrinarySediment Artifacts Artifact Comments Appearance Starch Granule • Resemble fat droplets when polarized • Dimpled center • From cornstarch used as glove powder • Highly refractile sphere Oil Droplets • From OIO contamination • highly refractile and may resemble RBCs Air Bubbles • occur when the specimen isplaced under a cover slip t
- 85. UrinarySediment Artifacts Artifact Comments Appearance Pollen grains • Concentric circles • spheres with a cell wall and occasional concentric circles Hairand fibers fromclothing and diapers • mistaken for casts • usually much longer and more refractile Fecal Artifacts • variety of sizes and shapes • presence of a fistula between the intestinal and urinary tracts • appear as plant and meat fibers or as brown amorphous material
- 86. END