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  1. 1. MICROSCOPIC URINALYSIS URINARY SEDIMENTSImportance  integral part of the urinalysis (UA).  Sediment findings often are necessary for the proper interpretation of results of the physicochemical portion of the UA  Detect and identify insoluble materials in the urine.Limitations  Least standardized  Time consuming  Expensive  Requires Technical expertise“ The decision to perform microscopic examinations should be made by each individual laboratory basedon its specific patient population.” -CLSISpecimen consideration  Fresh or adequately preserved  Mid-stream clean catch  First morning specimen  Thoroughly mixedVOLUME:  10-15 mL  Indicate if lesser volume is used  Correct for volumesSpecimen Centrifugation  5 minutes  400 RCF (relative centrifugal force)  Braking mechanism is not recommendedSEDIMENT PREPARATION  0.5 – 1.0 mL  Aspirate off the supernatant  Thorough resuspension of the sedimentSEDIMENT VOLUME:  20 uL (0.02 mL)  Cover slip overflow of sediment not allowedRCF  Centrifugal force – how many times greater than gravity- expressed as relative centrifugal force (RCF) or g
  2. 2. - depends on three variablesi. speed- expressed as revolutions per minute (rpm) **related to RCF by the following equation: RCF = 1.118 x 10-5 x r x (rpm)2ii. massiii. radius(r)- measured from the center of the centrifugal axis to the bottom of the test tube shield.Constituents of urine sediment  Organized sediment – biological source  RBC  WBC  Epithelial cells  Fats  Casts  Bacteria  Yeast  Fungi  Parasite  spermatozoa  Unorganized sediment chemical source  Normal acid crystals  Normal Alkaline crystals  Abnormal crystals of metabolic origin  Abnormal crystals of iantrogenic originRed Blood Cells (RBC)  Smooth, non-nucleated, biconcave disk  7 um in diameter  Examined under HPO  Reported as average in 10 HPFsVariations: 1. Crenated – found in concentrated urine 2. Ghost cells – cell membrane found in dilute urine 3. Dysmorphic – cells of varying sizes, have cellular protrusions or are fragmentedNormal and Crenated RBC
  3. 3. Normal and Dysmorphic RBCClinical Significance  Damage to the glomerular membrane  Vascular injury within the genito-urinary tract  Increased following strenuous exercise  Contamination with menstrual bloodGROSS HEMATURIA:  Advanced glomerular damage  Trauma damaging vascular integrity  Acute inflammation and infection  Coagulation disorderWhite Blood Cells (Neutrophils)  Larger than RBCs measuring 12 um in diameter.  Predominantly neutrophils  Contains granules and multi-lobed nuclei  Reported as average number per 10 hpf  “Glitter Cells” – disintegrated neutrophils  Easily lyses in dilute alkaline urine
  4. 4. Uncommon WBCEosinophils – associated with drug induced interstitial nephritis, UTI and renal transplant rejection  Reported as a percentage in 100 to 500 WBC  1 % eosinophil is significant  Preferred stain – HanselMononuclear Cells – increased in the early stages of transplant rejection  Appears vacuolated and contains inclusions  Diagnosed by cytocentrifugation and Wright’s stainClinical SignificancePyuria – increase in urinary WBCs  Indicates: -infection or inflammation in the genitourinary system -Bacterial infection: pyelonephritis, cystitis, prostatitis and urethritis -Non-bacterial disorders – glumerulonephritis, LE, Interstitial nephritis and tumorsEpithelial Cells  Represents the normal sloughing of old cells  Clinically insignificant in small numbers  May be contamination from the genitalia
  5. 5.  Three types are seen in urine classified according to their site of origin in the genitourinary tract. - Squamous epithelial cells - Transitional epithelial (Urothelial) cells - Renal Tubular Epithelial CellsSquamous epithelial cells  Largest cells found in the urine sediment  Contains abundant cytoplasm with a prominent nucleus  Originates from the linings of the vagina and female urethra and the lower portion of the male urethra.  No clinical significance  Usually increased in females  Reported in words as rare, few, moderate or manyClue Cells  Indicates infection vaginal infection with Gardnerella vaginalis  Squamous cells covered with the bacteria
  6. 6. Transitional Epithelial Cells(Urothelial)  Smaller than squamous cells  Appears in several forms (polyhedral, spherical, caudate) due to its ability to absorb water.  Originates from the lining of the renal pelvis, calyces, ureters and bladder and the upper portion of the male urethra  No clinical Significance  Increased in invasive urologic procedures such as catheterization  Presence of vacuoles and irregular nuclei may indicate viral infection or malignancyRenal Tubular Epithelial Cells  The most clinically significant of the epithelial cells.  Morphology varies depending on the site of origin PCT – largest of the RTE, rectangular, coarsely granular cytoplasm. DCT – smaller, round or oval, with eccentrically placed round nucleus Collecting ducts – cuboidal, never round, eccentrically placed nucleus, one side is straight, appearsin sheets
  7. 7. RTE Clinical Significance  Increase indicate necrosis of the renal tubules - exposure to heavy metals - drug induced toxicity - hemoglobin and myoglobin toxicity - viral infection -pyelonephritis - allergic reaction - malignant infiltration -salicylate poisoning -acute allogenic transplant rejectionOval Fat Bodies  RTE cells that absorb lipids present in the glomerular filtrate  Highly refractile  Seen along with free-floating fat droplets  Stains well with Sudan III and Oil Red O  Composed of triglycerides, neutral fats and cholesterol  “Maltese Cross” – observed in the presence of cholesterol under polarized light
  8. 8. Maltese CrossCasts Unique to the kidney Most difficult to recognize and most important sediment Represent a biopsy of the tubules Must be observed under subdued light because of the low refractive index of the cast matrix Reported as the average number in 10 lpfs Disintegrates in dilute alkaline urine.Composition of Casts Major constituent: Tamm-Horsfall protein The glycoprotein gels easily under conditions of urine stasis, acidity and the presence of sodium and calcium Width of the cast depends on the size of the tubuleCast Formation Formed in the lumen of the DCT and collecting ducts
  9. 9. Hyaline Cast The most commonly seen cast in the urine Consist almost entirely of Tamm-Horsfall protein Colorless, homogenous, non-refractive, semi-transparent 0-2/lpf is normal Seen in strenous exercise, dehydration, heat exposure, and emotional stress Increased in acute glomerulonephritis, pyelonephritis, chronic renal disease and congestive heart failure
  10. 10. RBC Cast Indicates bleeding within the nephron Primarily associated with damage to the glomerulus. Also associated with proteinuria and dysmorphic RBC Orange-red in color Dirty brown cast indicates hemoglobin degradation and associated with acute tubular necrosis.
  11. 11. WBC Casts Indicates infection or inflammation within the nephron. Associated with pyelonephritis and differentiates upper UTI from lower UTI Also seen in acute interstitial nephritis and glomerulonephritis. Appears granular and multilobed
  12. 12. Epithelial Cell Casts Contains RTE cells Indicates advanced tubular destruction Seen in heavy-metal and drug induced toxicity, viral infections, allograft rejections and pyelonephritis.Fatty Casts Associated with oval fat bodies and free fat droplets in cases of lipiduria Indicates nephrotic syndrome, toxic tubular necrosis, DM and crush injuries Highly refractile, confirmed with Sudan III and Oil Red O using polarized light
  13. 13. Mixed Cellular Casts Contains more than one type of cell Usual combinations: WBC and RBC in glomerulonephritis, WBC and RTE or WBC and bacteria in pyelonephritisMakes identification difficultGranular Casts May appear finely of coarsely Non-pathologic increase in strenuous exercise In diseases, it indicates disintegration of cellular casts
  14. 14. Waxy Casts  Represents extreme urine stasis indicating chronic renal failure.  Presents brittle, highly refractile cast matrix due to disintegration of hyaline and other cellular components of the cast  Appears fragmented with jagged edges and notches on their sides
  15. 15. Broad Casts Also referred to as renal failure casts Represents extreme urine stasis Indicates destruction of the tubular walls Commonly of the granular and waxy types.
  16. 16. Cast FormationOther Casts Rare incorporation of other structures in the urine sediments Pigmented Casts – hemoglobin, myoglobin and certain drugs Hemosiderin casts Crystal casts – urates, calcium oxalates and sulfonamidesCylindroids Resemble casts but have one end that tapers to a tail Found in conjunction with casts and have same significance
  17. 17. Mucus Threads  Long thin waxy threads, very transparent  Can be found in small number in normal urine  Increased numbers indicate inflammation or irritation of the urinary tract Bacteria  Not normally seen in urine  Results from vaginal, urethral, external genitalia or specimen container contamination  Presents as cocci or bacilli  Usually motile  May Indicate UTI if seen in freshly voided urine and correlated with WBCsYeast  Small, refractile, oval structures which may show budding  In severe in fections, mycelium may be seen  Most common: Candida albicans
  18. 18.  Seen in DM, immunocompromised patients and women with vaginal moniliasis Accompanied by WBCsFungi In severe infections May include appearance of myceliumParasites Most frequent: Trichomonas vaginalis – pear shaped flagellate with undulating membrane In fresh wet preparations, usually motile with rapid darting movements
  19. 19.  Other parasites: Schistosoma haematobium, Enterobius vermicularis, other parasite contaminants from the fecesT. vaginalisS. haematobiumE. VermicularisSpermatozoa
  20. 20.  Oval, slightly tapered heads and long flagella like tails, usually non-motile Seen in urine of both female and male after intercourse and in male urine after masturbation and nocturnal emission Not clinically significant except in cases of male infertility and retrograde ejaculation Also important in medico-legal casesUrinary Crystals Formed by the precipitation of urine solutes Rarely of clinical significance Reported in words Identified in order to detect the few abnormal crystalsCrystal Formation In vivo factors include: the concentration and solubility of crystallogenic substances contained in the specimen, the urine pH the excretion of diagnostic and therapeutic agents. In vitro factors include: temperature (solubility decreases with temperature), evaporation (increases solute concentration), urine pH (changes with standing and bacterial overgrowth).Amorphous Urates Amorphous urates appear as aggregates of finely granular material without any defining shape Amorphous urates (Na, K, Mg, or Ca salts) tend to form in acidic urine May have a yellow or yellow-brown color. Common in refrigerated specimens wit pink sediments
  21. 21. Uric Acid  May appears as Rhombic, foursided flat planes, wedges, and rosettes  Usually yellow-brown but may appear colorless  Highly birefrigent under polarized light  Increased in high levels of purines and nucleic acids  Seen in patients with leukemia undergoing chemotherapy, Lesch-Nyhan syndrome and gout Calcium Oxalate Dihydrate  Calcium oxalate dihydrate crystals typically are seen as colorless squares whose corners are connected by intersecting lines (resembling an envelope).  They can occur in urine of any pH.  The crystals vary in size from quite large to very small.  In some cases, large numbers of tiny oxalates may appear as amorphous unless examined at high magnification.
  22. 22.  Increased in high intake of oxalic acid and ascorbic acidCalcium Oxalate Monohydrate Less frequently seen Oval or dumbbell shaped Birefrigent Indicates ethylene glycol poisoningNormal Crystals seen in neutral to alkaline urine Amorphous phosphate Triple Phosphate Calcium Phosphate Calcium Carbonate Ammonium BiurateAmorphous Phosphates Morphologically resemble amorphous urates Increased in refrigerated sample but gives a white color Can be differentiated from urates by the pH of the urine and its non-dissolution on warming.
  23. 23. Triple Phosphate, Struvite,Ammonium Magnesium phosphate appear as colorless, 3-dimensional, prism-like crystals ("coffin lids"). Occasionally, they instead resemble an old-fashioned double-edged razor blade Birefrigent on polarized light
  24. 24. Calcium Phosphate Colorless Shape: long, thin prisms with one pointed and arranged as rosettes or clusters of needles Thin irregular plates that float on surface of urine Associated with renal calculi Dissolves in dilute acetic acid  May be confused with sulfonamide crystal
  25. 25. Calcium Carbonate Calcium carbonate crystals usually appear as large yellow-brown or colorless spheroids with radial striations. They can also be seen as smaller crystals with round, ovoid, or dumbbell shapes Liberates gas on addition of acetic acidAmmonium Biurate Color: yellow to brown Shape: Spherical bodies with long irregular spicules Often described as thorn- apple Associated with the presence of ammonia from urea-splitting bacteria Soluble in acetic acid and HeatAbnormal Urine crystals of Metabolic Origin Seen in acidic to neutral urine Requires chemical confirmation - Cystine -Tyrosine
  26. 26. -Leucine -Cholesterol - BilirubinCystine Colorless, refractile, hexagonal plates that are often laminated Seen in patients with cystinuria Disintegrates in alkaline urine Soluble in ammonia and dilute HCl Confirmed by the cyanide nitroprusside reactionTyrosine Colorless, fine, silky needles arranged in sheaves or clumps Seen in hereditary tyrosinosis, oasthouse urine disease and with leucine in massive liver failure Confirmed by the nitrosonaphthol test or HPLC
  27. 27. Leucine Yellow, oily looking spheres with radial and concentric striations Extremely rare Seen in severe liver damage with tyrosine
  28. 28. Cholesterol Color: transparent Shape: regular to irregular flat plates with one corner notched out, may be single or in larger numbers Most often found after refrigeration Indicates Excessive tissue breakdown Seen in nephritis and nephritic syndrome Soluble in chloroformBilirubin Bilirubin crystals tend to precipitate onto other formed elements in the urine. fine needle-like crystals can form on an underlying cell. This is the most common appearance of bilirubin crystals.
  29. 29.  cylindrical bilirubin crystals can form in association with droplets of fat, resulting in a "flashlight" appearance. This form is less commonly seen. Seen in Obstructive jaundice Bilirubin must be present in urineAbnormal Crystals of Iantrogenic origin Caused by increased amount of drugs
  30. 30.  Important because of the likelihood of renal damage and bleeding leading to renal failure -Sulfonamides -Ampicillin -Radiographic contrast mediaSulfonamides Color: brown to yellow Shape: needle-like shapes seen in bundles or sheaves; Stacks of wheat Common forms: sulfamethoxazole, acetylsulfadiazine and sulfadiazineAmpicillin Long, thin, colorless needles in acidic urine Very rarely seen Seen in Administration of large parenteral doses
  31. 31. Radiographic contrast media  Color: opaque , appear dark and thick  Shape: pleomorphic needles, single or sheaves  May be mistaken for cholesterol crystals  Significant in elderly patients  Intravenous injection for radiography Can appear up to 3 days after injectionContaminants and Artifacts  Usually easy to see  Causes distraction on the observer - Starch -Fibers -Air bubbles
  32. 32. -Oil droplets-Glass Fragments- Stains- Pollen grains- Fecal contamination

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