1. Urine routine and microscopy
KEM Hospital,Mumbai.

2. Urinanalysis is fundamental test that should be performed in all urologic patients.
It will help in conveying the structural and functional status of urinary system and associated abnormalities in it.

3. Collection of urine specimen-
In the male patient, a midstream urine sample is obtained after retracting the prepuce.
In female before obtaining sample clean the vulva and separate the labia.Mid stream urine is collected.
To evaluate for a possible infection in a female, a catheterized urine sample should always be obtained.
In infants sterile plastic bag with adhesive collar over genitalia can be used.Suprapubic aspirate can be used.
Need to be examined within 1 hour

4. Clean Catch

5. Routine urine analysis includes-
Physical examination
Chemical examination
Microscopic examination

6. Physical examination Urine volume Colour Turbidity PH
Specific gravity (SG) & Osmolality

7. Urine dipstick analysis-
Glucose
Bilirubin
Ketones
Specific Gravity
Blood pH
Protein
Urobilinogen
Nitrite
Leukocyte Esterase

8. Colour-
Color : normally , pale to dark yellow (urochrome) Abnormal color : some drugs cause colour changes 1. red urine : causes: hematuria hemoglobinuria myoglobinuria 2. yellow-brown or green-brown urine: bilirubin cause : obstructive jaundice

9. Red Urine Microscopic Hematuria Urinary tract source
Pseudohematuria (non-hematuria related red urine)
Myoglobinuria
Hemoglobinuria
Phenolphthalein Laxatives
Phenothiazines
Porphyria
Rifampin
Pyridium
Bilirubinuria
Phenytoin
Foods (Beets, Blackberries, Rhubarb)
Microscopic Hematuria
Urinary tract source
Urethra or bladder
Prostate
Ureter or kidney
Non-Urinary tract source
Vagina
Anus or rectum

10. Appearance Clarity: normally, clear
Abnormal colour: cloudy urine/turbidity
Causes: 1. crystals or nonpathologic salts
phosphate, carbonate in alkaline urine
(dissolve---add acetic acid)
uric acid in acid urine
(dissolve---warming to 60℃)
2. various cellular elements: leukocytes,
RBCs, epithelial cells
3.Chyluria,Lipiduria
4.Hyperoxaluria,Hyperuricosuria

11. Odour has little diagnostic significance.
Aromatic odour------> Normal urine due to aromatic acids.
Ammonia odour------> On standing due to decomposition of urea.
Fruity odour > Diabetes due to the presence of ketones.
An infection with E. coli bacteria can cause a bad odor

12. Urine volume The average adult : 1000ml to 2000ml/24h Increase
Polyuria---more than 2000ml of urine in 24 hr
1. physiological states: water intake, some
drugs, intravenous solutions
2. pathologic states: diabetes mellitus,
diabetes insipidus

13. Urine volume Decrease Oliguria---less than 400ml of urine in 24 hours
Anuria---less than 100ml of urine in 24 hours
1. prerenal: hemorrhage, dehydration,
congestive heart failure
2. postrenal: obstruction of the urinary tract
(may be stones, carcinoma)
3. renal parenchymal disease:
acute tubular necrosis, chronic renal failure

14. Specific gravity (SG) Reflect the density of the urine
Ranges from to 1.035
Increase(>1.020): Dehydration、Decreased fluid intake,Diabetes mellitus,SIADH.
Decrease(<1.008): Increased water intake,diabetes insipidus,diuretics,decreased renal concentrating ability.
Fixed specific gravity of is seen in renal insufficiency either acute or chronic.
Urine osmolality ranges from 50 to 1200 mosm/lit

15. The Urine Dipstick: Specific Gravity 1.000 Chemical Principle 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:

16. Urine PH- Normal PH- Ranges from 4.5 -8
The average ranges from 5.5 to 6.5
Higher PH---alkaline urine
1.drugs: sodium bicarbonate
2.Renal tubular acidosis type I
3.alkalosis (metabolic or respiratory)
Lower PH---acid urine
1.drugs: ammonium chloride
2. acidosis (metabolic or respiratory)

17. Methyl Red (at high concentration; low pH) and
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.:

18. Chemical examination Protein Glucose Ketones Occult blood Bilirubin
Urobilinogen
Nitrites & Leukocyte esterase.

19. The Urine Dipstick: Protein Chemical Principle
“Protein Error of Indicators Method”
Negative
Trace
+ (30 mg/dL)
++ (100 mg/dL)
+++ (300 mg/dL)
++++ (2000 mg/dL) Pr H
Tetrabromphenol Blue
(buffered to pH 3.0) H+ Pr Pr Pr Pr Pr Pr
Read at 60 seconds
RR: Negative

20. Proteins in urine
Normal adult excrete less than 150mg of protein in urine within 24 hours.
Urine proteins come from plasma protein(albumin,globulins) and Tamm-Horsfall (T-H) glycoprotein .
More than 150mg proteins in urine in 24 hours is called as proteinuria.
It can be quantified as-
heavy proteinuria----＞4.0g/24 hours
moderate proteinuria g/24 hours
minimal proteinuria----＜1.0g/24 hours

21. Pathophysiological causes-
1)Glomerular proteinuria:
damage to glomerular basement membrane but tubular function is normal
It can be selective proteinuria-chiefly albumin or nonselective proteinuria
heavy proteinuria
acute glomerulonephritis,Diabetes

22. 2)Tubular proteinuria-
-Renal tubular disease damage tubular function but glomerular function is normal
-Moderate proteinuria
- disease: pyelonephritis
3)Overflow proteinuria-
Excess levels of a protein in the circulation,hemoglobin, myoglobin, etc. The renal function is normal
Hemoglobinuria
Myoglobinuria
Multiple Myeloma
Amyloidosis

23. Glucose in urine- Normally all glucose filtered is absorbed in PCT.
Renal threashold for serum glucose is 180mg/dl
Glycosuria-
1.hyperglycemia: diabetes mellitus
Cushing’s syndrom
2.without hyperglycemia: renal tubular
dysfunction such as pyelonephritis

24. Horseradish Peroxidase
Chemical Principle
Negative
Trace (100 mg/dL)
+ (250 mg/dL)
++ (500 mg/dL)
+++ (1000 mg/dL)
++++ (2000+ mg/dL)
Glucose Oxidase
Glucose + 2 H2O + O2 --->
Gluconic Acid + 2 H2O2
Horseradish Peroxidase
3 H2O2 + KI ---> KIO3 + 3 H2O
Read at 30 seconds
RR: Negative

25. Limitations of Urine Glucose Detection
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
CuSO4 test for reducing sugars.

26. Acetoacetic Acid + Nitroprusside ------> Colored Complex
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

27. Uses and Limitations of Urine Ketone Detection
Normally not seen in urine.
Significance
- Diabetic ketoacidosis
- Prolonged fasting
-pregnancy
-rapid weight loss
Limitations
- Interference: expired reagents (degradation with exposure to moisture in air)
- Only measures acetoacetate not other ketone bodies(acetone, beta hydroxy butyric acid)

28. The Urine Dipstick: Bilirrubin Negative Chemical Principle + (weak)
++ (moderate)
+++ (strong)
Chemical Principle
Acidic
Bilirubin + Diazo salt > Azobilirubin
Read at 30 seconds
RR: Negative

29. Urobilinogen + Diethylaminobenzaldehyde -------> Colored Complex
The Urine Dipstick:
Urobilinogen
0.2 mg/dL
1 mg/dL
2 mg/dL
4 mg/dL
8 mg/dL
Chemical Principle
Urobilinogen + Diethylaminobenzaldehyde
> Colored Complex
(Ehrlich’s Reagent)
Read at 60 seconds
RR: mg/dL

30. Significance
- Increased direct bilirubin seen in obstructive jaundice.
-Increased urobilinogen is seen in hemolysis and hepatocellular diseases.
Limitations
- Interference: prolonged exposure of sample to light
- Only measures direct bilirubin--will not pick up indirect bilirubin
- Serum test for total and direct bilirubin is more informative

31. The Urine Dipstick: Blood Negative Trace (non-hemolyzed)
Chemical Principle
Trace (non-hemolyzed)
Lysing agent to lyse red blood cells
Moderate (non-hemolyzed)
Diisopropylbenzene dihydroperoxide +
Tetramethylbenzidine
> Colored Complex
Trace (hemolyzed)
Heme
+ (weak)
++ (moderate)
Read at 60 seconds
RR: Negative
Analytic Sensitivity: 10 RBCs
+++ (strong)

32. 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

33. Types of hematuria-
1)Glomerular-dysmorphic RBCs with cast and proteinuria.
e.g.-Glomerulonephritis all types.
2)Non glomerular-Normal RBCs no cast,proteinuria+
e.g.-tubulointerstitial diseases,renovascular HTN
3)Essential/surgical-Normal RBCs no cast,no proteinuria
e.g.Tumors,stones,UTI

34. The Urine Dipstick: Nitrite Chemical Principle Negative Positive
Acidic
Negative
Positive
Nitrite + p-arsenilic acid > Diazo compound
Diazo compound + Tetrahydrobenzoquinolinol
> Colored Complex
Read at 60 seconds
RR: Negative

35. Uses and Limitations of Nitrite Detection
Significance
- Gram negative bacteriuria(convert nitrate to nitrites)
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

36. The Urine Dipstick: Leukocyte Esterase Chemical Principle
Derivatized pyrrole amino acid ester
> 3-hydroxy-5-phenyl pyrrole
Negative
Trace
+ (weak)
++ (moderate)
+++ (strong)
Esterases
3-hydroxy-5-phenyl pyrrole + diazo salt
> Colored Complex
Read at 2 minutes
RR: Negative
Analytic Sensitivity: 3-5 WBCs

37. Indicates presence of WBCs in urine. Significance
Leukocyte Esterase Detection
Indicates presence of WBCs in urine.
Significance
- Pyuria
- Acute inflammation
- Renal calculus
Both leucocyte esterase test and nitrite test used in conjuction to screen for UTI.
If these tests are positive then microscopic analysis of urine should be done.

38. Microscopic Examination
General Aspects
Preservation
- Cells and casts begin to disintegrate in hrs. at room temp.
- Refrigeration for up to 48 hours (little loss of cells).
Specimen concentration
- Ten to twenty-fold concentration by centrifugation.(3000 rpm for 5 minutes of ml urine sample)
-Urinary sediment should be examined with both low power(100x) and high power field(400X)
-Low power- RBC,WBC,cast,cystine crystal,trichomonas,schistosoma.
-High power- Dysmorphic RBCs,other crystals,bacteria,yeast.

39. Types of microscopy - Phase contrast microscopy - Polarized microscopy
- Bright field microscopy with special staining
(e.g., Sternheimer-Malbin stain)

40. Microscopic Examination
Abnormal Findings
Per High Power Field (HPF) (400x)
> 3 erythrocytes
> 5 leukocytes
> 2 renal tubular cells
> 5 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

41. Microscopic Examination
Bacteria & Yeasts
Bacteria
- Bacteriuria-More than 5 per HPF reflects CFU 1,00,000/ml
Yeasts
- Candidiasis-Most likely a contaminant but should correlate with clinical picture.
Viruses
CMV inclusions-Probable viral cystitis.
Parasites-
-Trichomonas,Schistosoma

42. Urinary casts-
Urinary casts are cylindrical structures produced by the
kidney and present in the urine in certain disease states.
They are protein coagulum formed in the distal convoluted tubule (DCT) and collecting ducts of nephrons and traps any luminal contents within matrix then dislodge and pass into the urine, where they can detected by microscopy.
-Urinary casts may be made up of cells (such as white
blood cells, red blood cells, kidney cells) or substances
such as Tamm-Horsfall mucoprotein.

43. Microscopic Examination
Casts
Erythrocyte Casts:Glomerular diseases
Leukocyte Casts:Pyuria, glomerular disease
Hyaline casts are composed primarily of a mucoprotein (Tamm-Horsfall proteins) secreted by tubule cells.
Granular cast or waxy cast arise from further degeneration of cellular elements.
Fatty cast- Nephrotic syndrome

44. Hyaline Casts appear Transparent

45. Red Cell Casts 45

46. White Cell Casts 46

47. Crystals in urine Normal-Cholesterol Crystals in acidic urine
Uric acid
Calcium oxalate
Cystine
Leucine
Crystals in alkaline urine
Ammonium magnesium phosphates(triple phosphate crystals)
Calcium carbonate

48. Calcium Oxalate dihydrate Crystals-Envelope shaped

49. Calcium Oxalate monohydrate Crystals-Dumbell shaped
Dumbbell Shape

50. Triple Phosphate Crystals-coffin lid shaped.

51. Urate Crystals-rhomboid

52. Microscopic Examination
Leucine Crystals

53. Cystine Crystals-Hexagonal

54. Ammonium Biurate Crystals
Microscopic Examination
Ammonium Biurate Crystals

55. Microscopic Examination
Cholesterol Crystals

56. Summary-Normal values
Color – Yellow (light/pale to dark/deep amber)
Clarity/turbidity – Clear or cloudy
pH – 4.5-8
Specific gravity –
Glucose -Negative
Ketones – None
Nitrites – Negative
Leukocyte esterase – Negative
Bilirubin – Negative
Urobilirubin – Small amount (0.5-1 mg/dL)
Blood - ≤3 RBCs
Protein - ≤150 mg/d
RBCs - ≤3RBCs/hpf
WBCs - ≤2-5 WBCs/hpf
Squamous epithelial cells - ≤15-20 squamous epithelial cells/hpf
Casts – 0-5 hyaline casts/lpf
Crystals – Occasionally
Bacteria – None

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