Urine examination-physical and chemical

1. Functions Of Kidney
I. Elimination Of Metabolic Waste Products.
II. Fluid , Electrolytes & Acid-base Status
Regulation.
III. Hormonal Regulation With Erythropoietin &
Renin Production ,Vit.D Activation.

2. Routine Urine Examination
PHYSICAL
EXAMINATION
CHEMICAL
EXAMINATION
MICROSCOPIC
EXAMINATION

3. I. Physical Examination
1) Total Volume
Normal Volume:600-2000ml/Day
a) Polyuria:urine Volume >2000ml/Day
 Causes :-Osmotic Diuresis
:-Polydipsia(diabetes Insipidus)
:-Diuretics
:-Chronic Renal Failure

4. • Oliguria
b) EXCRETION <400ML/DAY
 CAUSES :-DEHYDRATION
:-CHF
:-SHOCK
:-RENAL ARTERY EMBOLISM
:-ACUTE GLOMERULONEPHRITIS
:-ATN
:-RENAL FAILURE
:-GU TRACK OBSTRUCTION

5. C) ANURIA :- EXCRETION <100ML/DAY
Causes :-Bilateral Complete Urinary
Tract Obstruction
:-Acute Cortical Necrosis
:-Necrotizing
Glomerulonephritis

6. 2) Specific Gravity: It Reflects The Degree Of
Concentration Of Urine.
Methods:
1)reagent Strips
2)refractometer
3)urinometer
4)falling Drop Method

7. 1.Reagent Strips
 Indirect Method
 Reagent Area Contains 3 Main
Ingredients:1)polyelectrolytes
2)indicator Substance
3)buffer
 PRINCIPLE:IT IS BASED ON THE Pka CHANGE OF THE
PRE-TREATED POLYELECTROLYTES IN RELATION TO THE
IONIC CONC.Of Urine.When The Ionic Conc.IS HIGH
,Pka IS DECREASED AS IS THE Ph.The Indicator
Substance Then Change Color Relative To The Ionic
Conc & This Is Translated To The Specific Gravity
Values.
 Advantage:not Affected By High Amounts Of
Glucose,protein Or Radiographic Contrast Media.

8. 2)refractometer
 Principle:refractive Index Of A Solution Is
Related To The Content Of Dissolved Solids
Present.
 It Is Temperature Compensated Hand
Model(60 To 100f)
 Requires Only Few Drops Of Urine.

9. 3)URINOMETER
IT IS A HYDROMETER ADOPTED
TO DIRECTLY MEASURE THE
SPECIFIC GRAVITY OF URINE AT
ROOM TEMP.

10. 4)falling Drop Method
 Direct Method
 More Accurate Than Refractometer
 More Precise Than Urinometer

11. Interpretation:
Normal-1.016 To 1.022/24 Hr
Increased In:-proteinuria
:-Diabetes Melitus
:-Diuretics
:-Antibiotics
:-Acute Glomerulonephritis
Decreased In:-renal Insufficiency

12. Diurnal Variation:higher In The Morning
&Lower In Afternoon
Fixed Specific Gravity:sp.GRAVITY OF ANY
SAMPLE OF URINE COLLECTED REMAINS FIXED
E.G.1.010
Seen In Chronic Glomerulonephritis

13. 3.Color
COLOR CONDITIONS
PALE YELLOW NORMAL URINE
RED Hb
ANILNE DYES
PHENOPHTHELIN
SMOKY RED OR BROWN BLOOD
DRUGS LIKE LEVODOPA
YELLOWISH ACRIFLAVIN
YELLOWISH GREEN BILE PIGMENTS
CAROTENE
ACRIFLAVIN
YELLOW BROWN BILIRUBIN
BROWN BLACK HOMOGENTISIC ACID
MELANIN
METHEMOGLOBIN
PORTWINE PHORPHYRINS

14. Physical Examination
4) Appearnce
• Normal:clear
• Milky:lipiduria
:Chyluria
:PRESENCE OF Pmns
• Cloudy:phosphates
:Carbonates
:Uric Acid
:Bacteria
:Sperm

15. Physical Examination
5)odour
• Normal:urinoid
• Maple Syrup-maple Syrup Urine Disease
• Mousy-phenylketonuria
• Sweet-ketosis
• Fishy-tyrosinemia
• Sulfurous-cystinuria
• Lack Of Odour- ARF(due To ATN)

16. Chemical Examination
1)proteins
Commonly Perfomed Methods Are:
I) REAGENT STRIPS:
MECHANISM:THIS METHOD TAKES ADVANTAGE
OF THE PROTEIN ERROR OF Ph INDICATORS.
PROTEINS CARRY A CHARGE AT
PHYSIOLOGICAL Ph , THEIR PRESENCE WILL
ELICIT A Ph CHANGE.

17. Chemical Examination
-In The Absence Of Protein The Strip Is Yellow.
-30-60 Sec. Following Urine Application, Variable
Shades Of Green Depending On The Type And
Conc. Of Protein Present.
-Results Read In A “Plus” System As:
Negative
Trace
1+ ( 30 Mg% )
2+ ( 100 Mg%)
3+ (300mg%)
4+ (>2gm%)

18. Chemical Examination
Ii) HEAT & ACETIC ACID TEST
• Principle: Proteins Are Precipitated When
Boiled In Acid Solution.
• Method:
• -Fill A Test Tube With Urine Up To 2/3.
• -Boil The Upper Portion By Holding The Tube
Up At The Bottom

19. Chemical Examination
-Appearance Of Cloudiness Or Precipitates On
Top;may Be Due To Proteins Or Phosphates.
-Add 1 T0 3 Drops Of 10%acetic Acid & Boil The
Top Portion Again.
-Cloudiness Due To Phosphates Will Disappear
But Not Due To Proteins.

20. Chemical Examination
Iii) SULPOSALICYLIC ACID TEST
-Mix Equal Amounts Of Urine With 5% Salphosalicylic Acid In A Test
Tube.
-Grading
Negative: No Turbidity(5mg/Dl Or Less)
TRACE: PERCEPTIBLE Turbidity(20mg/Dl)
1+ :Distinct Turbidity,no Discrete
Granulation(50mg/Dl)
2+ :Turbidity With Granulation,no
Flocculation(200mg/Dl)
3+ :Turbidity With Granulation And
Flocculation(500mg/Dl)
4+ :Clumps Of Precipitated Protein Or Solid Precipitate
(1gm/Dl OR MORE)

21.  Causes Of Proteinuria
• It Is Divided In To:
1)pre-renal:-chf
-Cerebral Injury
-Severe Infections
-Fever
2)renal:-glomerulonephritis
-Nephrosclerosis
-Diabetes Glomerulosclerosis
-Nephrotic Syndrome
-Pyelonephritis
3)post-renal:-inflammation Of The
Pelvis Of Kidney, Ureter, Bladder

22. Quantification Of Proteinuria
• HEAVY Proteinuria(>4gm/Day)
- Nephrotic Syndrome Associated With
1 Primary Renal Disease:
Idiopathic
ARPGN
Ch. Gn
2 Systemic Disease With Renal Involvement
Dm
Sickle Cell Disease
Sle
3 Others
Severe CHF
Renal Vein Thrombosis
Malignant HT

23. Quantification Of Proteinuria
• MODERATE PROTEINURIA (1 To 4 Gm/Day)
- Nephrosclerosis
- Multiple Myeloma
- Toxic Nephropathy
- Presence Of Calculi
• MINIMAL Proteinuria(<1gm/Day)
- Chronic Pyelonephritis
- Chronic Interstitial Nephritis
- Congenital- Polycystic Kidney Disease
- Medullary Cystic Disease

24. Qualitative Proteinuria
• It Requires Electrophoretic Separation Of Urine
Proteins.
1. Glomerular Pattern:
Less Selective Proteinuria Suggest Severe Glomerular
Damage.
2. Tubular Pattern:
Loss Of LMW Proteins E.G. B2microglobulin
Light Chain Ig
Tubular Proteinuria Seen In:
Fanconi’s Syndrome
Cystinosis
Wilson’s Disease
Pyelonephritis

25. Qualitative Proteinuria
3. Overflow Proteinuria
- Due To The Overflow Of Excess Levels Of A
Protein In The Circulation.
- Initially Not Associated With Glomerular Or
Tubular Damage But Themselves Cause Renal
Damage E.G. Myoglobin Causing ATN

26. Qualitative Proteinuria
4.Bence- Jones Proteinuria
- It Represents Either Kappa Or Lambda Ig Light
Chain.
• Causes
-Multiple Myeloma
-Monoclonal Gammopathy
-Cryoglobulinemia
-Primary Amyloidosis
-Adult Fanconi’s Syndrome

27. 1) Heat Method
-To 10 Ml Of Fresh Clear Urine ,Add 2ml Of
Saturated Sod. Chloride Soln To Prevent
Precipitation Of Mucin.
-Add Acetic Acid Soln Drop By Drop To Make Ph
Slightly Acidic.
-Heat The Urine In A Beaker Or Waterbath
Controlling The Temp.With A Thermometer.

28. -If B-J Proeins Are Present ,The Urine Becomes
Cloudy Between 40 & 50C And A Flocculant
Precipitate Appears To 50c.
-On Raising The Temp.To 100c The Precipitate
Dissolves Completely Or Partially & Reappears
On Cooling.

29. Qualitative Proteinuria
2) Electrophoresis
-Presence Of B-J Globoulin Is Indicated By Single
Sharp Peak In The Globulin Region On
Electrophoresis.
-More Specific Than Heat Method.
-Positive Test For B-j Proteins By Heat Method
Should Always Be Confirmed By
Electrophoresis

30. Qualitative Proteinuria
5. Micro Albuminuria
- Presence Of Albumin In Urine Above The Normal
Level But Below The Detectable Range Of
Conventional Urine Dip Stick Method.
- Measured By
1 Immunological Method
2 Nephelometric Method
3 Radioimmuno Assay
*Causes:
- Indicator Of Early & Possibly Reversible
Glomerular Damage
- Dm
- Ht

31. Chemical Examination
 Othe Types Of Proteinuria
1)postural Proteinuria:first Morning Urine Before Arising
Shows High Specific Gravity But No Protein.Protein
Only Appears After Person Is Upright.
-Usually <1.5gm/Day
-Benign Condition
-Slowly Disappears With Time
-Occurs In 15% Of Apparently Healthy Persons
-SEEN IN SOME Pts. With Resolving Acute
Pyelonephritis Or Gn.

32. Chemical Examination
2)transient Proteinuria:commonly Found In Routine Urinalysis
Of Asymptomatic Healthy Children & Young Adults Initially
Progressive Renal Disease Is Not Present.
Usually < 2 Gm/Day
Disappear With Recovery From Precipitating Cause.
-Associated With High Fever
-Chf
-Hypertension
-Stress
-Exposure To Cold
-Strenous Exercise
-Seizures

33. Chemical Examination
3)persistent Proteinuria
A)glomerular :
IDIOPATHIC
o Membrenoproliferative Gn
o Membrenous Glomerulopathy
o Minimal Change Disease
o Focal Segmental Glomerulosclerosis
o Amyloidosis

34. Chemical Examination
 Secondary
o Infections
o Vascular
o Drugs(nsaids,heroin,gold,penicillamine
o Autoimmune
o Neoplasia
o Hereditary & Metabolic Ds.
B)decreased Tubular Reabsorption
 Acquired
o Drugs
o Heavy Metals
o Sarcoidosis
o Acute Tubular Necrosis

35. Chemical Examination
o Interstitial Nephritis
o Acute & Chronic Pyelonephritis
o Renal Graft Rejection
 Congenital
o Fanconi Syndrome
o Oculo-cerebral-renal Syndrome
 Hereditary
o Wilson Ds.
o Sickle Cell Ds.
o Medullary Cystic Ds.

36. Chemical Examination
2)sugar & Other Reducing Substances
Methods
I.Reagent Strips
-Based On A Specific Glucose Oxidase & Peroxidase Method
-Specific For Glucose
-Doesn’t React With Other Reducing Substances
• Interpretation
-Negative
-Trace ( 100mg%)
-1+ (250 Mg%)
-2+ (500mg%)
-3+ (1 Gm% )
-4+ (> 2 Gm%)

38.  False Positive Results
-Exposed To Air
-Peroxidase Contamination
-Oxidising Agents
 False Negative Results
-ASCORBIC ACID >25mg%
-Drugs(aspirin)
-Specific Gravity >1.020
-HIGH Ph

39. Chemical Examination
Ii. Benedict’s Qualitative Test
Principle:cupric Ions Are Reduced To Cuprous
Form By Sugar & Other Reducing Sustances When
Heated.
Method :
1] To 5ml Of Qualitative Benedict’s Reagent,8
Drops Of Urine Are Added & Mixture Is Boiled For
2 Minutes
2] Examine After Cooling

40. Chemical Examination
 Interpretation :Test Is Positive If The Originally Blue
Reagent Is Discoloured With The Formation Of
Precipitate; Olive Green,yellowish,orange Or Brick
Red(depending Upon The Amount Of The Reducing
Substances Present)
 Sugars Reducing The Benedict’s Reagent:
-Glucose
-Fructose
-Galactose
-Pentoses

41. Chemical Examination
Other Reducing Substances :
-Homogentisic Acid
-Gluconates
-Creatinine
-Uric Acid
-Certain Drugs

42. Chemical Examination
 Causes
1)hereditary
I. Galactosemia
II. Galactokinase Deficiency
III. Sever Liver Disease With Galactose Intolerance
IV. Hereditary Fructose Intolerance
V. Phenylketonuria
2) Due To Hyperglycemia
I. Diabetes Melitus
II. Cushing’s Syndrome
III. Administration Of Hormones
IV. Drugs(morphine,anesthetic Drugs)

43. Chemical Examination
3)renal
I. Tubular Origin(s.Glucose<180mg%,oral & INTRAVENOUS
GLUCOSE TOLERANCE TESTS ARE NORMAL,KETOSIS IS
ABSENT)
II. Fanconi Syndrome
III. Toxic Renal Tubular Acidosis
IV. Inflammatory Renal Disease
V. Glomerular Due To Increased Glomerular Filtration Rate
Without Tubular Damage)
4)lactosuria
-During Lactation & Late In Normal Pregnancy
-Sepsis

44. Chemical Examination
3)ketone Bodies
Acetone,aceto-acetic Acid & 3-hydroxybutyrate Are 3
Ketonbodies Seen In Urine.
Methods
I.Reagent Strips
-Based On A Nitroprusside Reaction For Ketones
1)chemistrip
-Detects About 10mg /Dl Of Acetoacetic Acid & 70mg/Dl
Of Acetone
2)multistix
-Detects 5-10mg/Dl Of Aceto-acetic Acid
-Doesn’t React With Acetone

45. Chemical Examination
Ii.Rothera’s Test For Acetone & Aceto-acetic Acid
Method
-Take 5 Ml Of Urine In A Test Tube
-Add Approx.1gm OF AMMONIUM SULPHATE & 3
DROPS OF FRESHLY PREPARED SODIUM
NITROPRUSSIDE SOLN.
-Now Run Along The Side Of The Tube,liquor
Ammonia So As To Form A Layer On The Top.
-If Permangenate Colour Ring Appears At The
Junction Of The Two Fluid,test Is Positive.

46. Chemical Examination
3)gerhardt’s Test For Acetoacetic Acid
-To 10 Ml Of Fresh Urine ,Add A Few Drops Of
10%ferric Chloride Drop By Drop Until A
Precipitate Is Formed.
-Filter The Precipitate & To Filtrate Add More
Ferric Chloride Drop By Drop .
-If Test Is Positive,a Violet Red Colour Develops.

47. Causes Of Ketonuria
-Metabolic Conditions
E.G.Dm
Renal Glycosuria
-Dietary Conditions
E.G.Starvation
High Fat Diets
-Increased Metabolic Requirements
E.G.Hyperthyroidism
Fever

48. Chemical Examination
4)bile Salts & Bile Pigments
Methods:
A)gmelin’s Test For Bile Pigments
-Take 3 Ml Of Conc. Nitric Acid In A Test Tube.
-Place Equal Amount Of Urine On It.
-Shake The Tube Gently From Side To Side.
-Note The Color Change.
-Interpretation: If Bile Pigments Are Present ,
There Is Play Of Colors: Yellow,red,violet,blue &
Green.

49. B) Harrison Fouchet’s Test For Bile Pigments
- 10 Ml Urine + 5 Ml Bacl2 Soln.
-It Gives Precipitates.
-Filter It Through Whatman No.2 Filter Paper.
-Gently Transfer Ppt. On Other Filter Paper.
-Now Add 1 To 2 Drops Of Fouchet’s Reagent
On It.
* Interpretation : It Gives Green Or Blue Color
If Bilirubin Is Present.

50. c) Hay’s Sulphur Flower Test For Bile Salts
-Take 5 Ml Of Urine In A Large Test Tube Or
Beaker.
-Sprinkle Sulphur Powder On The Surface.
* Interpretation : If Bile Salts Are Present,
Sulphur Particles Sink To Bottom Of The Tube.
o Inreased In Jaundice.

51. Microscopic Examination
• 15ml Of Mixed Sample Of Urine Is Spun At
2000rpm For 10mins.
• The Sediment Is Suspended In 1 To 2 Drops Of
Urine & Coverslip Preparation Is Made For
Microscopy.
• It Is Performed To Detect Cells,casts &
Crystals.

52. Microscopic Examination
1}cells
A)red Blood Cells
-Pale Biconcave Discs.
Normal: 0 To 2 Cells/Hpf
Abnormal: >3 Cells/Hpf
Causes: Trauma
Gn
Intestitial Nephritis
Calculus
Tumors
Acute Infection
Tb
Atn

53. • DYSMORPHIC Rbcs
-Cells With Protusions Or Fragmentation
-Their Presence Strongly Suggests Renal
Glomerular Bleeding.
B)leucocytes
I)neutrophils:granular Sphere About 12um With
Multilobulated Nuclei.
Normal:<5 Leucocytes /Hpf
Females May Show Higher Number

54. • Pyuria-increased No.Of Leucocytes In Urine
Causes:pyelonephritis
Cystitis
Prostatitis
Urethritis
Acute Urethral Syndrome
Fever
Physiological-following Exercise

55. C)eosinophils
-Not Normally Seen
-Seen In -Tubuloiterstitial Ds Associated With
Hypersensitivity To Drugse.G.Penicillin
-Disorder Of GUT
-Uti
-Renal Transplant Rejection
D)lymphocytes & Mononuclear Leucocytes
-Small Lymphocytes Are Normally +Nt In Urine
-Mononuclear Cells>30% Suggests Chronic
Inflammation

56. E)epithelial Cells
I)squamous Epi.Cells
-Seen In Normal Urine
Ii)transitional Epi.Cells

57. Iii)renal Tubular Epi.Cells
-Small No.+Nt Normally
-Increased In-atn
-Drugs
-Heavy Metals

58. Iv)collecting Duct Cells
-Increased In :
-Renal Transplant
Rejection
-Atn
-Ischemic Injuries
-Malignant Nephrosclerosis
-Acute GN With Tubular Damage
-Drugs & Chemicals

59. • 2}casts
-These Are Cylindrical Structures With Parallel Edges,
Basically These Are Precipitates Of Proteins Formed In
DCT And Collecting Tubules.
-Tamm-horsfall Protein,constituting About 1/3 Of Total
Urinary Protein, Forms Matrix Of All Casts.
-Classification Of Casts
I.Matrix
Ii.Cellular
Iii.Inclusions
Iv.Pigments

60. I.Matrix Casts
1)hyaline Casts
-0 To 2 Casts/Lpf Is Normal
-Increased In-renal Ds.
-Transiently Increased In-exercise
-Heat Exposure
-Dehydration
-Fever
-Chf
-Diuretic Therapy

61. 2)waxy Casts
Seen In
-Tubular Inflammation
& Damage
-Crf
-Acute & Chronic Renal Allograft Rejection

62. Ii.Cellular Casts
1)rbcs CASTS
Seen In:agn
Ig Nephropathy
Lupus Nephritis
Sabe,infarction

63. 2)leucocyte Casts
Seen In-tubulointerstitial Ds
-Glomerular Ds.
-Interstitial Nephritis.
-Lupus Nephritis

64. 3)renal Tubular Epithelial Cell Cast
Seen In
-ATN
-Viral Ds E.G.Cmv
-Exposure To Drugs
-Heavy Metal Poisoning
-Allograft Rejection
4)mixed Cells
-Erythrocytes,neutrophils & Renal Tubular Cells

65. Iii.Inclusions
1)granular Cast
Seen In
-Glomerular & Tubular Ds
-Tubulointerstitial Ds.
-Renal Allograft Rejection
-Pyelonephritis
-Viral Infections
-Chronic Lead Poisoning

66. 2)fatty Casts
Seen In Nephrotic Syndrome
3)crystal Cast
-Cast Containing Urate,ca Oxlate &
Sulfonamides.
4)haemosiderin Granules
5)melanin Granules

67. Iv.Pigmented Casts
1.Hb Cast
Seen With -Erythrocyte Cast
-Glomerular Ds.
2.Myoglobin Cast
Seen In Myoglobinuria
Arf
3.Bilirubin & Other Drug Casts
Seen In Obstructive Jaundice.
V.Broad Casts
Seen In CRF
Indicates Tubular Dilatation

68. 3}crystals
I.Crystals In Acidic Urine
A) Ca.Oxlate Crystals
-Oxaluria
-Crohn’s Ds.
-Oxalate Stones
-Methoxyfurane Toxicity

69. B) Amorphous Urate
- Precipitate On Standing In Conc. Urine
C) Crystalline Uric Acid
Seen In Pt. On Chemotherapy
Lesch-nyhan Syndrome
Uric Acid Stones

70. Ii.Crystals Found In Alkaline Urine
A)triple Phosphate
B)amorphous
Phosphate
C)ca Carbonate
D)ammonium Biuret
- All Are Seen In UTI

71. Iii. Other Crystals In Urine
A)cysteine Crystals
-Cystinuria
-Cystine Calculi

72. B)tyrosine Crystals
-Severe Liver Disease
C)leucine Crystals
-Severe Liver Disease
D)sulfadiazine Crystals
-Pt On Sulfonamide Therapy
E)ampicillin (High Dose)
F)radiographic Media
-After I.V. Radiographic Studies

73. 4) Abnormal Cells & Other Formed Elements
I)tumor Cells
-Exfoliated From Renal Pelvis,ureter,bladder
Wall & Urethra
Ii)viral Inclusion Cells
Herpes –Synctial Giant Cells Containing
Eosinophilic ,Intranuclear Inclusions
CMV -Enlarged Cells With Basophilic
Intranuclear Inclusions
Polyoma –Dense,basophilic ,Homogenous
Intanuclear Inclusions

74. Iii) Platelets
Up To 30,000/Ul Seen In HUS
Iv)bacteria
Significant When >100000 Organisms/Ml With
Gram’s Stain
V)fungi
Candidia
Yeast
Vi) Parasites
Commonly Found Parasites Are
-Trichomonas Vaginalis
-Schistosoma Haematobium
-Entamoeba Histolytica

75. Reference Values For The Urinary
Sediment Considered As Abnormal
• Erythrocytes>5 Cells At Hpf
• Leukocytes>5 Cells At Hpf
• Renal Tubular Cells>2 Cells At Hpf
• Transitional Cells>5 Cells At Hpf
• Squamous Cells Rarely Significant
• Hyaline Casts>3 Cast At Lpf
• Granular Casts>1 Cast At Lpf
• Pathological Casts Any
• Crystals Any

76. Special Examination
1) Quantitative Sugar Estimation
-By Quantitative Benedict’s Test
-Having Prognostic Value In DM
-But Now A Days Not Useful Because
Of Availability Of Glycosylated Hb.

77. 2) Quantitative Protein Estimation
-For This 24 Hrs Urine Collection Sample Is
Required
A)esbach’s ALBUMINOMETER
-Urine Is Filled Up To “U” Mark.
-Esbach’s Reagent Is Filled Up To “R” Mark.
-Keep It Overnight.
-Next Day Precipitated Protein Form Whitish Clot
Like Coagulant Will Be Read As Per Marking On
Instrument.
-Result Is Recorded In Gm/Lit/24 Hrs.

78. • B) Biuret Method
– Colorimetric Method
– Less Time Consuming
– More Specific
– Routinely Used And More Reliable
 PRINCIPLE :PEPTIDES OF PROTEIN REACT WITH ALKALINE
COPPER TARTARATE SOLN.TO GIVE A VOILET COLOURED
COMPLEX WHICH IS MEASURED COLORIMRTRICALLY AT
540nm WHICH IS DIRECTLY PROPORTIONAL TO THE
CONC.Of Protein In Specimen.
C)sulphosalicylic Acid /Tca Method
– Turbidimetric & Semi-quantitative Methods

79. 3)urobilinogen
-Normally Present In Urine.
-Unstable In Acid Urine
-When Exposed To Light,reduced To Form A Pigment
Urobilin,hence Urine Sample Is Collected In Amber
Coloured Bottle.
Method
Principle:formation Of Red Coloured Azo Dye With Diazonium
Compound
-To 10ml Of Urine,add 1ml Of Ehrlich’s Reagent
-If Intense Red Colour Develops,further Testing Is Done By
Using Serial Dilution Of Urine .
-Reading Is Recorded Up To The Test Tube Showing Faint Pink
Colour.

80. NORMAL: 1:10 To 1:30 Dilution
Significance:
*Hemolytic Jaundice-increased
*Hepatocellular Jaundice-early Stage->increased
Late Stage->decreased
*Obstructive Jaundice-absent Or Decreased

81. Biochemical Tests Perfomed In Blood
 Blood Urea
 Serum Creatinine

82.  Blood Urea
• End Product Of Proteins & Amino Acid
Metabolism
• Produced By Liver
• Excreted Mainly By Kidneys
• Measured By Following Methods

83. I.Berthelot Method
Principle:urea Is Converted To Ammonium By
The Use Of Urease.Ammonium Ion Then
Reacts With A Mixture Of Salicylate,sodium
Nitroprusside & Hypochlorite To Yield A Blue –
Green Chromophore.The Intensity Of The
Color Formed Is Proportional To The Urea
Concentration In The Sample.
It Is Colorimetric End Point Test.

84. Reagents
Reagent I : Urea Enzyme Reagent
Reagent II : Urea Color Developer
Urea Standard : 50mg/Dl
Sample
Serum From Plain Bulb
Plasma From Edta,citrate Bulb

86. BLANK STANDARD SAMPLE
REAGENT I 1 ml 1 ml 1 ml
STANDARD - 10ul -
SAMPLE - - 10ul
MIX WELL & INCUBATE FOR 5 mins AT 37c OR 10 mins AT RT
REAGENT II 1 ml 1 ml 1 ml
MIX WELL & INCUBATE FOR 5 mins AT 37c OR 10 mins AT RT

87. • Measure The Absorbance Of Standard & Test
Against Reagent Blank At 578nm.
• Calculation
• Urea ( Mg/Dl)=(at-ab/AS-AB) X Conc.Of Std.
• Normal Value: 15 – 40 Mg/Dl
• Bun=urea X (M.W. Of Nitrogen /M.W.Of Urea)
• Normal Value: 5 – 20 Mg/Dl

88. Other Methods
Ii.Nessler Method
Principle : When Diluted Serum Is Incubated With
Urease Powder ,It Produces Liquid
Ammonia.After Depolarization With
Zn(oh)2,color Is Produced By Ammonia With
Nessler’s Reagent,which Is Read At Green Filter.
Iii.Oxime Method
Principle : On Heating Diacetyle Monoxime Give
Hydroxylamine & Diacetyl Which React With Urea
& Give Color Complex.

89. • Limitation Of Urea Measurement
1)considerable Glomerular Damage Must Occur
Before Urea Increases.
2)urea Depends On :
i) Protein Intake In Diet
ii) Catabolic State Of Protein
iii) High Protein Load E.G.Upper G.I.Bleeding
iv) Advanced Liver Disease
v) State Of Hydration

90. AZOTEMIA: Biochemical Abnormality
Denoting Higher Level Of BUN In Blood.
Uremia :Clinical Menifestation Comprised Of
Increased Urea In Plasma,acidemia
,Electrolyte Imbalance Associated With Clinical
Features Such As Vomiting,nausea,anaemia,
Altered Mentation & Altered Hemostasis

91.  Increased Level Of Urea Seen In :
I)increased Tissue Protein Catabolism
Seen In-fever
-Diabetic Coma
-Thyrotoxicosis
-After A Major Operation
Ii)excess Brekdown Of Blood Protein
Seen In -Leukemia
-Gastrointestinal Ds.
Iii)diminished Excretion Of Urea
-Commonest & Most Common Cause
-Divided Into

92. 1)pre-renal
-Urine Is Hypertonic
-Mild Proteinuria
-Urine Sediment Contain Hyaline Or Granular Casts
Due To Decreased Renal Blood Flow:
-Shock
-Chf
-Dehydration
-Haemorrhage
-Renal Artery Stenosis

93. 2)renal
Due To Decreased Glomerular Filtration
-Arf
-Crf
-Gn
-Pyelonephritis
-Tubular Necrosis
-Interstitial Nephritis

94. 3)post Renal
Due To Obstruction In Urinary Tact
-Ureteric Stone
-Urethral Stone
-Urethral Stricture
-Bladder Stone
-Prostatic Hypertrophy
-Prostatic Tumor

95.  Serum Creatinine
• Creatinine Is Endogenous Substance Produced By
The Muscle From Creatine & Creatine Phosphate
By A Non-enzymatic Dehydration Process.
• Most Widely Used As Marker Of Gfr Because
I)endogenous Substance Of Fairly Constant Rate Of
Production
Ii)not Bound To Plasma Proteins ,Therefore Is
Filtered Freely By The Glomerulus
Iii)not Reabsorbed By The Renal Tubules & Only A
Small Amount Is Secreted By The Tubules.

96. I. Alkaline Picrate Method
PRINCIPLE : In An Alkaline Medium ,Creatinine
Reacts With Picrate To Produce An Orange Yellow
Colour Which Is Proportionaal To The Creatinine
Concentration In Sample.
Reagents :
1.Picric Acid
2.Sodium Hydroxide
3.Creatinine Standard- 2mg/Dl
Sample :
Serum Or Plasma

99.  Procedure
-Reading Is Taken At 510nm
TEST STANDARD BLANK
PICRIC ACID
0.5% NaOH
PICRIC ACID
0.5% NaOH
DISTILLED
WATER
PICRIC ACID
0.5%NaOH
DISTILLED
WATER
TEST STANDARD BLANK
PICRIC ACID 500ul 500ul 500ul
0.5%NaOH 500ul 500ul 500ul
DISTILLED
WATER
- - 50ul
STANDARD - 50ul -
SERUM 50ul - -

100. Normal Values
Male :0.6- 1.2 Mg%
Female :0.5-1.0 Mg%
Other Methods
Ii.ENZYMATIC METHOD : Creatinine Is Hydrolysed
By Creatinine Iminohydrolase To Ammonia & N-
methylhydantoin.The Ammonia Then Combines
With 2-oxoglutarate & NADH In The Presence Of
Glutamate Dehydrogenase To Produce Glutamate
& NAD+.The Consumption Of Nadh,measured As
Decreased In Absorption At 340nm.

101. Iii.High Perfomance Liquid Chromatography
-Excellent Specificity
-Requires Deproteinization Of Sample
-Lengthy Procedure

102. • Creatinine Vaues Increase In
1.Skeletal Muscle Causes
-Trauma
-Rapidly Progressive Muscular Dystrophy
-Poliomyositis
-Dermatomyositis
-Myesthenia Gravis
-Starvation
-Sepsis
-Major Surgery

103. 2.Acute & Chronic Renal Diseases
In Renal Ds.Creatinine Tends To Rise Slowly
Than Urea.
3.Others
-Diabetic Acidosis
-Purperium

104. Bun : Creatinine Ratio
• Helps To Differentiate Pre-renal & Post-renal
Azotemia From Renal Azotemia.
• Normal Value : 12 To 16
1.Increased Ratio With Normal Creatinine
A)pre Renal Azotemia(due To Decreased GFR)
B)catabolic States With Increased Tissue
Breakdown
C)gi Hemorrhage

105. D)high Protein Intake
E)impaired Renal Function Plus-
-Excess Protein Intake Or Tissue Breakdown
-Urine Reabsorption (E.G.Ureterocolostomy)
-Patients With Reduced Muscle Mass
F)selective Increase In Plasma Urea During Use
Of Loop Diuretics
2.Increased Ratio With Elevated Creatinine
-Post Renal Azotemia E.G.Obstuctive Uropathy
-Pre Renal Azotemia Superimposed On Renal
Disease

106. 3.Decreased Ratio With Decreased Bun
-Atn
-Low Protein Diet
-Severe Liver Ds.
-Repeated Dialysis
-Inherited Deficiency Of Urea Cycle Enzymes
-Pregnancy
4.Decreased Ratio With Increased Creatinine
-Rhabdomyolysis
-Phenacemide Therapy
-Muscular Patients Developing Renal Failure

107. Tests Measuring Gfr
• GFR Is Measured By Clearance Tests.
• Clearance Test Gives Relatively Accurate &
Useful Measure Of The Gfr & Also The
Excretory Capacity Of The Kidney.
Substances Used For Measuring Gfr :
I. Exogenous Substances
I)inulin
Ii)mannitol
Iii)sodium Thiosulphate
Iv)tc-dtpa( Diethylene Triamine Penta-acetic

108. Ii.Endogenous Substances
I) Creatinine
Ii) Urea
-It Requires 24 Hrs Urine Collection

109. Equation For Clearance Is :
Cx=uxv/Px
Cx=clearance Of A
Substance X
Ux=conc.Of The Substance X
In Urine
Px=conc.Of Substance X In
Plasma
V=volume Of Urine Per Unit
Time

110. Interpretation
o Good Estimate In Pt.With Reduced GFR.
o Inaccurate If S.Creatinine Is Not Stable
o Unreliable In Pts.With Extremes Of Age,body
Size Or Composition
o A Normal GFR In Association With Impaired
Concentrating Ability May Be Found In Sickle
Cell Anaemia,diabetes
Insipidus,pyelonephritis.
o Increased Gfr Is A Risk Factor For Progressive
Nephropathy In Dm

111. o If Urea Clearance Value Falls
<20% - Severe Renal Failure
<5% - Uremic Coma

112. STAGE DESCRIPTION GFR (ml/min/1.73m2)
STAGE 1 KIDNEY DAMAGE WITH
NORMAL OR INCREASED
GFR
>90
STAGE 2 KIDNEY DAMAGE WITH
MILD DECREASE IN GFR
60 - 89
STAGE 3 MODERATE DECREASE IN
GFR
30 - 59
STAGE 4 SEVER DECREASE IN GFR 15 - 29
STAGE 5 RENAL FAILURE < 15 OR DIALYSIS

113. Determination Of Acute Renal
Failure
• To Differntiate Pre-renal Azotemia & Acute
Tubular Necrosis.
• Fractional Excretion : Is The Quantity Of A
Substance Excreted In Urine Expressed As A
Fraction Of The Filtered Load Of The Same
Substance.
Fe(sod.) =( Usod./Psod.) X (Pcreat/Ucreat)
• If It Is < 0.01 – Pre Renal Azotemia
• If It Is >0.01 - Atn

114. Urine & Serum Diagnostic Indices
PRE RENAL RENAL POST RENAL
URINE SODIUM <20 mEq/ L >20 mEq/ L >20 mEq /L
URINE CHLORIDE <20 mEq/L >20 mEq /L --------
FENa < 1 % > 2% >2%
FEUrea < 35% ------ ------
URINE
OSMOLARITY
>500 mg% < 350 mg% < 350 mg%
URINE/SERUM
CREATININE
> 40 < 20 < 20
URINE/SERUM
UREA
> 8 < 3 < 3
RENAL FAILURE
INDEX
< 1% > 1% >1%
BUN : CREATININE > 20 ~ 10 ~10

115. Renal Biopsy
• Should Be Preceded By
I)confirmation That Two Kidneys Are Present
Ii)no Renal Infection Is Present
Iii)there Is No Bleeding Disorder
• Examination Should Include
I)histology –Stained By H &E,trichome,pas,silver
Ii)immunofluorescence-with Antisera Specific For
Igg,iga,igm,c1q,c3,c4,fibrinogen,albumin,kappa &
Lambda Light Chains
Iii)electronmicroscopy –Alport Syndrome ,Thin
Basement Membrane Nephropathy

116. • Indications For Biopsy
I)acute Renal Allograft Dysfunction
Ii)persistent Or Reccurent Haematuria With
Proteinuria
Iii)nephrotic Syndrome To Distinguish Etiology
Iv)proteinuria > 1gm/Day Or With Abnormal Urine
Sediment
V)nephrotic Syndrome- If Unresponsive To Therapy
Or Before Therapy
Vi)non-nephrotic Proteinuria With Progressive
Disease
Vii)evaluation Of Collagen Ds.(Sle)

117. • Contraindications For Biopsy
I)haemorrhagic Ds
Ii)solitary Kidney
Iii)active Kidney Infection
Iv)renal Artery Vasculitis With Aneurysm
V)hydronephrosis
Vi)uncontrolled Severe HT
Vii)uncooperative Pt.

118. References
Henry’s Clinical Diagnosis & Management By
Laboratory Methods
Interpretation Of Diagnostic Tests,jacques
Wallach.
Textbook Of Medical Laboratory
Technology,godkar
Www.Google.Com