urine 1.pptx RTS455Y5S656XXTYS56SE4EEDT FFHR6

TT67D67

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2. Composition of normal urine (24 hrs) in adults 2 Sl No Parameters Values Sl No Parameters Values 1 Volume 600-2000 ml 10 Urea nitrogen 12-20 gm 2 Specific gravity 1.003-1.030 11 U r i c A c i d 250-750 mg 3 Osmolality 300-900 mOsm/kg 12 Sodium 40-200 mEq 4 pH 4.6-8.0 13 Potassium 25-125 mEq 5 Glucose < 0.5 gm 14 Chloride 110-250 mEq 6 Proteins < 150 mg 15 Calcium (low cal c i u m d i et) 50-150 mg 7 Urobilinogen 0.5-4.0 gm 16 Formiminogluta mi c ac i d (FIGlu ) < 3 mg 8 Prophobilinog en 0.2 mg 17 Red cells, epithelial cells and w hi t e blood cells ≤ 1-2 per high p o w er f ie l d 9 Creatinine M: 14-26 mg/kg F: 11-20 mg/kg

3. Indication of urine analysis 3 Suspected renal diseases Glomerulonephritis Nephrotic Syndrome Pyelonephritis Renal failure Urine tract infection diagnosis Metabolic disorders i.e. Diabetes mellitus Differential diagnosis of jaundice Detection of Plasma cell dyscrasias Diagnosis of pregnancy

4. Collection of urine: Time of collection (1/3) Single specimen First morning voiding M ost c o n c e n t r a t ed Acidic pH Formed element preserved like casts, cells Used for R outine e x ami na t ion Fasting Glucose Protein Nitrite Pregnancy test M i c r os c opic a n a ly sis of c ell u lar ele m e n ts Orthostatic proteinuria Bacteriological analysis 4

5. Collection of urine: Time of collection (2/3) Single specimen The Random Specimen Can be collected at any point of time Routine examination Post-prandial specimen C olle c t ed t w o ho u r s af t er a m eal i n th e af t ern o on Insulin therapy monitoring in Diabetes Mellitus Urobilinogen 5

6. Collection of urine: Time of collection (3/3) 24 –hour-specimen First urine discarded in morning Clean 2 litres bottle with cap- used Whole day and night urine collected Next day first urine in morning also collected Preserved at 4-6 ◦ C during collection After collection immediate transportation to lab Thoroughly mixed and a p a r t of w hole sam p le use d f or qu a ntitat i v e estimation of Protein Hormones 6

7. Collection of urine: Methods of collection 7 Midst r eam specimen U sed f or all ty p es of e x amination C ollec t ed af t er v oiding in i tia l half of urine Clean-catch specimen Used for bacteriological culture Urethral opening- cleaned with soap and water C ollec t ed af t er v oiding in i tia l half of urine Cathe t er specimen Used for bacteriological culture For bed ridden and obstructed urinary tract patient Infants Aspiration done above the symphysis pubis

8. Changes occur in room temperature after prolonged standing 8 Increase in pH- production of ammonia Formation of crystal- calcium and phosphate precipitation Loss of ketone bodies- volatile Decrease of glucose- glycolysis by bacteria and cells Oxidation of bilirubin to biliverdin Oxidation of urobilinogen to urobilin Bacterial proliferation Disintegration of cellular elements

9. Preservation of urine sample 9 T est t o b e don e within 2 hrs Can be kept at 4-6 ◦ C for maximum 8 hrs Routine analysis- preservatives avoided Preservative used in 24 hrs sample Hydrochloric acid- Adrenaline, Nor-adrenaline, vanillyl mandelic acid, steroids Toluene- measurement of chemicals Boric acid- general preservative Thymol- inhibit bacteria and fungi Formalin- formed elements

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11. Physical examination: appearance 11 Appearance Diagnosis Cause Whi t e and cl o u d y on standing in alkaline urine Disappear on a d ditio n of a d r op of dilu t e a c et i c acid Amorphous phosphates Pink and cloudy in acid urine Disso l v e on warming Amorphous urates Varying grades of turbidity Microscopy Pus cells U ni f orm l y cl o u d y , do not settle at the bottom following centrifugation Microscopy, nitrite test Bacteria

12. Physical examination: volume 12 24 hrs average urine output- 600 to 2000 ml (adult) Polyuria M o r e tha n 2 00 m l per d a y Occurs in Diabetes Mellitus, Diabetes Insipidus , Chronic renal failure, diuretic therapy. Oliguria L ess tha n 400 ml p er d a y Febrile states, acute glomerulonephritis, congestive cardiac failure, dehydration Anuria L ess tha n 100 m l p er d a y Acute tubular necrosis, acute glomerulonephritis, complete urinary tract obstruction

13. Physical examination: colour 13 Dif f e r ent c olours of ur i ne Found in colorless D i lu t e urine(diabe t es mel l itu s , diabetes insipidus, red Haematuria , hemoglobinuria, porphyria, myoglobinuria Dark brown to black Alkaptonuria, melanoma brown hemoglobinuria yellow Concentrated urine Y ell o w g r een or g r een biliverdin Deep y ell o w w i t h y ell o w f o a m bilirubin O r an g e or o r an g e b r o wn Urobilinogen, porphobilinogen Mil k y w hi t e chyluria R ed or o r an g e f luo r es c en c e w i th UV light porphyria

14. Physical examination: odour 14 Fruity: ketoacidosis, starvation, Musty: phenylketonuria Fishy: UTI ( Proteus ), tyrosinaemia Ammoniacal: UTI ( E. coli ) Foul: UTI Sulfurous: cystinuria

15. Physical examination : pH/reaction (1/2) Normal range: 4.6-8 Tested by Litmus paper pH indicator paper pH meter Reagent strip test 15

16. Physical examination: pH/reaction (2/2) 16 Acidic urine: Ketosis - diabetes mellitus, starvation, fever UTI by E. coli High protein diet Alkaline urine: St r ic t v e g eta r ian UTI by Proteus or Pseudomonas Se v er v o m i t ing CRF

17. Physical examination: Specific gravity (1/2) 17 Depe n d s on th e c on c e n t r ation of v arious solutes in the urine N ormal r an g e :1 . 3 t o 1 . 3 at 2 O C Measured by : Urinometer Refractometer Reagent strip method

18. Physical examination: Specific gravity (2/2) 18 High specific gravity Diabetes mellitus( glycosuria) Nephrotic syndrome (proteinuria) Fever Dehydration Low specific gravity Diabetes insipidus Chronic renal failure

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20. Chemical examination: Protein 20 Causes of proteinuria (1/3) Normal value: 150 mg per day Glomerul a r p r o t ein u ri a : du e t o in c r eased permeab i lity of g lomerular capilla r y w all Selective (only albumin and transferrin bands seen by electrophoresis) Nonselective (pattern same as serum)

21. Chemical examination: Protein 21 Causes of proteinuria (2/3) T ubu l ar p r o t e inuria: P r o t ei n uri a ca u sed b y r enal tubu l ar dysfunction. acute and chronic pyelonephritis, heavy metal poisoning, tuberculosis of kidney Overflow proteinuria: Proteinuria associated with increased production of abnormal low molecular weight proteins, Bence jones protein (plasma cell dyscrasias) Hemoglobin (intravascular haemolysis) Myoglobin (skeletal muscle trauma) Lysozyme (acute myeloid leukemia type M4 or M5)

22. Chemical examination: Protein 22 Causes of proteinuria (3/3) Hemodynamic proteinuria: alteration of blood flow causes increase protein filtration High fever Hypertension C on g est i v e ca r di ac fa i l u r e H e a v y e x e r cise Post-renal proteinuria: Inflammation and neoplasia of Renal pelvis Ureter Bladder Prostate Urethra

23. Tests for Detection of Protein: Heat and acetic acid test Principle: Protein get coagulated when boiled Method: 2/3 rd of test tube filled with urine Heat upper portion of urine in inclined test tube till it boils Cloudiness appears due to coagulation Few drops of 10% acetic acid added to dissolve phosphorus –helps to detect false positivity 23

24. Tests for Detection of Protein: Reagent strip method Principle: Indicator strip coated with bromo-phenol blue indicator Buffered to pH 3.0 with citrate Protein changes ionization of dye Semi quantitative method Notes: Mai n l y r e a ct i v e t o albu m in False negative- Bence Jones protein, myoglobin, haemoglobin False positive- Gross haematuria, highly alkaline urine, contaminated urine 24

25. Tests for Detection of Protein: Sulphosalicylic acid test Principle: Protein gets precipitated after adding organic acid like sulphosalicylic acid Method: 2ml of clear urine in a test tube If urine neutral or alkaline- add one drop of glacial acetic acid Add 2-3 drops of 3-5% sulphosalicylic acid Chec k tu r bidit y agai n st da r k backg r ou n d Notes: More sensitive and reliable than boiling test False positive- gross haematuria, highly concentrate urine, radiographic dye, excess uric acid, tolbutamide, sulphonamide, salicylates, penicillins F alse negat i v e - v e r y dilu t e urine Can detect- albumin, myoglobin, haemoglobin, Benes Jones P r o t ein 25

26. Test for detection of Glucose in urine 26 BENEDICT TEST (1/2) Principal: When urine is boiled in Benedict solution, blue alkaline copper sulphate is reduced to red brown cuprous oxide if a reducing agent is present. Method: Take 5ml of Benedict’s reagent in a test tube, add 8 drops of urine. Boil the mixture. Nil: no change from the Blue color Trace: Green without precipitate 1+(approx 0.5grams/dl)Green with precipitate 2+(approx 1.0 grams /dl) Brown precipitate 3+(approx 1.5 grams /dl) Yellow orange precipitate 4+(approx >2grams /dl ) Red precipitate

27. Test for detection of Glucose in urine BENEDIC T TE S T (2/ 2 ) Detects all reducing substances like glucose, fructose ,lactose Sensitivity is 200 mg of glucose /dl 27

28. Test for detection of Glucose in urine 28 R ea g ent strip method Spec i f i c f or g lu c ose Base d on g lu c ose o xid a se pe r o xid a se r eaction More sensitivity(sensitivity 100 mg glucose/dl)

29. Causes of glycosuria 29 G l y c osur i a with h ype r g l y caemi a : Diabetes Acromegaly C ushing ’ s dis e ase Hyperthyroidism Drugs like corticosteroids G l y c osur i a without h ype r g l y caemi a : R e n al tubula r d y sfunct ion

30. Ketone bodies 30 Types Acetone A c e t oa c e t i c acid β hydroxy butyric acid They are products of fat metabolism

31. Test for ketone bodies R othe r a ’ s t est Principal: A c e t o n e and a c e t o a c etic acid r eact w i t h sodium nit r o p rus s ide in the presence of alkali to produce purple color Method: Take 5ml of urine in a test tube and saturate it with ammonium sulphate, then add one crystal of sodium nitroprusside. then slowly run the liquor ammonia along the sides of the test tube. Formation of purple colored ring at junction indicates positive test R oth e r a ’ s t est i s se n sit i v e t o 1 -5 m g /dl of a c e t oa c e t a t e a n d 1 - 25 mg/dl of acetone 31

32. Other tests for detection of ketone bodies 32 Acetest tablet test F er r i c ch l oride t est (Ge r ha r dt ’ s t est) Reagent strip method

33. Causes of ketonuria 33 Diabetes N on diabet ic cause s : Hi g h f e v er Starvation Se v er v om i t i ng Diarrhoea G l y c o g en s t o r a g e diseases

34. Bi l irubin Fouchet’s test Principal: Bilirubin absorbs to Barium Chloride and results in green color formation when fouchet’s reagent is added. Method: In 5 ml of urine add 2.5 ml of 10% Barium chloride and mix well .then filter to obtain the precipitate on a filter paper .To precipitate add 1 drop of Fouchet’s reagent. De v elo p e m e n t of blu e g r e e n c olor a r ound th e d r op indicates presence of bilirubin 34

35. Other tests for bilirubin 35 Foam test Gmelin’s test Lugol’s iodine test R ea g e n t strips wi t h di a z o r ea g e n t

36. Urobilinogen Ehrlich test Method: In 5 m l of urin e add 0.5 m l of Eh r li c h ’ s r ea g e n t (HCL 20 ml ,distilled water 80 ml , para dimethylaminobenzaldehyde 2gm)Allow to stand for 5 mins. development of pink colour indicates positive test. 36

37. Causes of Urobilinogen in urine 37 Hemolytic jaundice Early hepatitis Hepatocellular jaundice

38. Bile salt 38 Hay’s surface tension test Method: In 5ml of urine sprinkle a pinch of sulphur p a rticle s . I f bil e salt i s p r esent s u lphur p a rticle s will sink to the bottom because bile salts lower the surface tension of urine .

39. Blood in urine 39 Benzidine test Principle : The peroxidase activity of haemoglobin decomposes hydrogen peroxide releasing nascent o x y g en , w hich i n t u r n o x i di z es ben z id i n e t o g i v e blue colour Method : Mix 2ml of benzidine solution with 2ml of hydrogen peroxide in a test tube .Take 2ml of urine and add 2ml of mixture .A blue or green colour within 5 min indicates positive reaction.

40. Causes of Haematuria 40 Disease of urinary tract Glomerular disease : Glomerulone p hritis Be r g e r ’ s disea s e Lupus nephritis Henoch-Schonlein purpura Non glomerular disease: Calculus , tumor, infection, tuberculosis, pyelonephritis, trauma, carcinoma of prostate Hematological condition: Coagulation disorders ,sickle cell disease

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42. Microscopic examination 42 Qualitative technique Urine must be freshly voided Examined without excessive delay in order to prevent cellular degeneration If preservative is required ,then 1 crystal of thymol or 1 drop of formalin (40%)is added to 10 ml of urine Well mix sample of urine(10 -15ml )is centrifuged in machine for 5mins at 1500 rpm. The top part(supernatant)is discarded. A drop of urine left at the bottom of test tube (sediment)is placed on the glass slide and c o v e r ed with c o v er slip i t i s e x amined un d er hi g h power

43. Red Blood cells Normally no RBC found Small, smooth, yellowish, a n uc l ea t e bi c onc a v e disks 7 µ in diameter- isomorphic Found Fresh urine 9-10 µ in diameter- swollen F ou n d i n h y p o t o n i c urine Smaller diame t er with spi k ey surface - crenated h y p er t o n i c urine Variable in size and shape- dysmorphic Glomerulonephritis 43

44. White Blood cells Spherical. G r anular with visible nuclei 10-15 µ in size distorted, smaller, and have f e w er g r a n ule s - degenerative WBC Infection – seen in clumps 44

45. Epithelial cells Small, polyhedral, c olu m ne r , or o v al h av e granular cytoplasm, e c c entric nucle u s se e n. Found in acute tubular necrosis , pyelonephritis, viral infection, 45

46. Urinary casts 46 Cylindrical aggregations of particles Form in the distal renal tubules and collecting ducts C omposed of a p r ecip i ta t io n of T am m - H orsfall p r o t ein

47. Types of urinary casts 47 A c ellular c asts H y aline casts G r anular casts W axy casts F a t t y casts Broad casts Cellular casts Red cell casts Whi t e c ell casts Epithelial cell casts

48. Hyaline casts Th e m ost c o mm on t ype of casts. Th ey a r e c y lin d rica l wi t h parallel sides and blunt rounded end, colourless, homogenous, transparent. Se e n i n f e v e r , st r e n ous exercise, glomerular proteinuria 48

49. Granular casts They are cylindrical s t ru c tu r e with c oarse or fine granules Seen in acute g lomer u lonephritis and pyelonephritis 49

50. Waxy casts They have homogenous smooth glassy appearance ,cracked or serrated margins and irregular broken off ends. Seen in end stage renal failure 50

51. Fatty casts They are cylindrical structures filled with highly refractile fat globules in Tamm Horsfall protein matrix Seen in nephrotic syndrome 51

52. Broad casts Form in dilated distal tubules Seen in chronic renal failu r e and se v er r enal t u b u l ar obs t ru c t i on Associated with poor prognosis 52

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54. Normal crystals in urine 54 Crystals present in acidic urine Uric acid crystals : These are variable in shape and are yellow brown in colour and soluble in alkali and insoluble in acid. F o u n d i n G o u t a n d leu k emia. Calcium oxalate crystals : These are colourless envelope-shaped .commonly found in diets rich in tomatoes, cabbages. Large number are seen in Ethylene glycol poisoning. Amorphous urates :These are urate salts of potassium, magne s ium , or calciu m i n acid urine Crystals present in alkaline urine Calcium carbonate crystals: small colorless, grouped in pair Ammonium phosphate crystals: yellow brown ,cactus like, called as thornapple Phosphates: Triple phosphates: colorless ,3-6 sided prisms with oblique surfaces at the end. Calcium hydrogen phosphate: colorless and variable shape Amorphous phosphate :colorless small granules.

55. Normal crystals found in urine 55 Calciu m o x ala t e T ri p le p hos p ha t e U ri c acid Calciu m ca r bo n a t e

56. Abnormal crystals in urine 56 Cysteine crystals: colorless clear hexagonal, highly refractile plates, soluble in hydrochloric acid. Seen in Cystenuria. Cholesterol crystals : colorless refractile, flat rectangular plates with notched corner. Seen in Nephrotic syndrome and hypercholesterolemia. Bilirubin crystal: brown and variable shapes. Seen in o b st r uct i v e l i v er disease. T y r osine c r y stals : clus t ers of f in e c olo r less or y ell o w needle s . Seen i n l i v er diseas e and T y r osinemia . Leucine crystals: refractile, yellow or brown spheres with radial or concentric striation. Seen in urine along with ty r osine i n se v er l i v er disease(c i r r hosis)

57. Abnormal crystals found in urine Cysteine Cholesterol B i l irubin Leucine 57

58. References 58 Burtis CA, Ashwood ER (Eds). Tietz fundamentals of clinical chemistry (5th Ed). Philadelphia; WB Saunders Company, 2001. Henry JB (Ed): Clinical diagnosis and management by laboratory methods. (20th Ed). Philadelphia; WB Saunders Company, 2001. Kawthalkar Shirish M, Essential of Clinical Pathology, Jaypee Brothers Medical Publishers (P) Ltd, ISBN: 978-93-80704-19-7, Pg no: 3-29 , India, 2010. World Health Organization. Manual of basic techniques for a health laboratory (2nd Ed). Geneva; World Health Organization, 2003.

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