3. MicroscopeMicroscope
Robert Hook – author of micrographia – primitiveRobert Hook – author of micrographia – primitive
microscope in 1665microscope in 1665
First compound microscope was based on theFirst compound microscope was based on the
advice of Kepleradvice of Kepler
Janssen and Janssen – first operationalJanssen and Janssen – first operational
microscope in 1950microscope in 1950
Eye piece magnification – 6x , 10x , 15xEye piece magnification – 6x , 10x , 15x
Condenser lens below the stage which focuses theCondenser lens below the stage which focuses the
light source on the specimen being observedlight source on the specimen being observed
Koehler illumination – even illumination across theKoehler illumination – even illumination across the
field of viewfield of view
4. Numerical aperture – ability of the lens to resolve whatNumerical aperture – ability of the lens to resolve what
is viewed , and just not to magnifyis viewed , and just not to magnify
NA = RI * sin aNA = RI * sin a
Dry objective NA 0.95Dry objective NA 0.95
Oil – NA = 1.4 is the bestOil – NA = 1.4 is the best
Low power – 2x and 4x – scanningLow power – 2x and 4x – scanning
Optical tube length – distance between the objectiveOptical tube length – distance between the objective
and image produced by the lensand image produced by the lens
For conventional microscopes the limit of usefulFor conventional microscopes the limit of useful
magnification is 1500xmagnification is 1500x
Resolution –measure of the power of a microscope toResolution –measure of the power of a microscope to
distinguish ,as separate entities ,two self illuminatingdistinguish ,as separate entities ,two self illuminating
points in close proximity to each otherpoints in close proximity to each other
5. Minimal resolvable distance =0.61 lambda /NAMinimal resolvable distance =0.61 lambda /NA
Spherical aberration – rays passing from peripherySpherical aberration – rays passing from periphery
will be brought to a smaller focal lengthwill be brought to a smaller focal length
Chromatic Aberration – refraction is greatest forChromatic Aberration – refraction is greatest for
violet light and smallest for red lightviolet light and smallest for red light
Apochromat lens – corrects both spherical andApochromat lens – corrects both spherical and
chromatic aberrationchromatic aberration
Plan apochromat lens – corrects both sphericalPlan apochromat lens – corrects both spherical
and chromatic aberration and gives a flat field ofand chromatic aberration and gives a flat field of
view – for critical photographyview – for critical photography
Achromat lens – cheapest..used for routineAchromat lens – cheapest..used for routine
purposespurposes
6. Koehler illuminationKoehler illumination
Light source – 12 inches from the microscopeLight source – 12 inches from the microscope
Hardened oil in objective – use a little XYLOL –Hardened oil in objective – use a little XYLOL –
don’t use muchdon’t use much dissolve the cement holding thedissolve the cement holding the
lenslens
Low power / high powerLow power / high power lower the condenserlower the condenser
and work with the concave mirrorand work with the concave mirror
Unstained specimen – close the diaphragmUnstained specimen – close the diaphragm
Oil immersion – raise the condenser up and useOil immersion – raise the condenser up and use
plane mirror – cedar wood oil / liquid paraffinplane mirror – cedar wood oil / liquid paraffin
Never use alcohol in any part of microscopeNever use alcohol in any part of microscope
7. Phase contrast microscopePhase contrast microscope
Further retardation of the indirect wavesFurther retardation of the indirect waves
which have passed through the cytoplasmwhich have passed through the cytoplasm
A halo appears around the objectA halo appears around the object
UsesUses
1.1. Unstained bacteriaUnstained bacteria
2.2. Amoeba , Trypanosomes , PromastigotesAmoeba , Trypanosomes , Promastigotes
3.3. Urine sedimentsUrine sediments
4.4. Platelets in suspensionPlatelets in suspension
9. Polarized light microscopePolarized light microscope
Birefringence – ability to pass light in a particularBirefringence – ability to pass light in a particular
plane – such materials are called anisotropicplane – such materials are called anisotropic
UseUse
1.1. Exogenous crystalline material – talc crystals atExogenous crystalline material – talc crystals at
injection sitesinjection sites
2.2. Endogenous Crystalline material – sodium urateEndogenous Crystalline material – sodium urate
(gout) , Calcium pyrophasphate(gout) , Calcium pyrophasphate
3.3. Collagen – dull yellow white colourCollagen – dull yellow white colour
4.4. Formalin - Heme pigment – artefact of poorFormalin - Heme pigment – artefact of poor
fixation – light micro – black stippled …polarizedfixation – light micro – black stippled …polarized
– whi– whi
5.5. tete
6.6. Amyloid stained with Congo red dyeAmyloid stained with Congo red dye
Oval fat bodies in urine gives a MALTOSEOval fat bodies in urine gives a MALTOSE
CROSS appearance in birefringenceCROSS appearance in birefringence
10. Interference microscopeInterference microscope
Light beam split – one beam enters –otherLight beam split – one beam enters –other
bypass—later combined –they interfere withbypass—later combined –they interfere with
each othereach other
We can find the DRY WEIGHT of the objectWe can find the DRY WEIGHT of the object
– because it is related to the refractory index– because it is related to the refractory index
Thickness of the object can also beThickness of the object can also be
determineddetermined
11. Electron MicroscopeElectron Microscope
Knoll & Ruska -1932Knoll & Ruska -1932
RPRP 15 – 30 A15 – 30 A
100x -50 000x100x -50 000x
Fixed with gluteraldehyde and embedded in hard plasticFixed with gluteraldehyde and embedded in hard plastic
materialmaterial
Stain – Osmium Tetroxide (+fixative)Stain – Osmium Tetroxide (+fixative)
UseUse
1.1. Happy mitochondriaHappy mitochondria
2.2. Premelanosomes in the cells of melanomaPremelanosomes in the cells of melanoma
3.3. Neurosecretory granulesNeurosecretory granules
4.4. Membrneous GNMembrneous GN
Osmium - fixes unsaturated lipids and phospholipids wellOsmium - fixes unsaturated lipids and phospholipids well
Gluteraldehyde – fixes proteins wellGluteraldehyde – fixes proteins well
Double fixation – gluteraldehyde as primary followed by postDouble fixation – gluteraldehyde as primary followed by post
– fixation with osmium tetroxide– fixation with osmium tetroxide
12. Resolving power of compound microscopeResolving power of compound microscope
is about 0.275 micronsis about 0.275 microns
Maximum limit of resolving power of humanMaximum limit of resolving power of human
eye is 25 micronseye is 25 microns
Thickness of specimen to study underThickness of specimen to study under
microscope is 1 -2 micronsmicroscope is 1 -2 microns
Fluorochrome – rhodamineFluorochrome – rhodamine
The general rule of thumb is that totalThe general rule of thumb is that total
magnification should not exceed 750 – 100magnification should not exceed 750 – 100
x the numerical aperurex the numerical aperure
14. Nephron is the organ unit for urine formationNephron is the organ unit for urine formation
William Bowman in 1842 – injection withWilliam Bowman in 1842 – injection with
potassium chromate and lead acetatepotassium chromate and lead acetate
GFR – 127ml/minGFR – 127ml/min
16 ml / min gain access to the DCT16 ml / min gain access to the DCT
Rate of urine formation = 1 ml / minRate of urine formation = 1 ml / min
180 L / day180 L / day
99% reabsorbed99% reabsorbed
Urine formed is only 1.5 – 1.8 L / dayUrine formed is only 1.5 – 1.8 L / day
Range = 0.5 -2.5 LRange = 0.5 -2.5 L
Specific gravity = 1.003 – 1.030Specific gravity = 1.003 – 1.030
Reaction is acidicReaction is acidic
Total solids present is 30 -70 g / LTotal solids present is 30 -70 g / L
15. ConstituentsConstituents Per 24 hrsPer 24 hrs
SodiumSodium 3 - 4 g3 - 4 g
CreatinineCreatinine 1 -1.8 g1 -1.8 g
PotassiumPotassium 1.5 - 2 g1.5 - 2 g
ChlorideChloride 9 - 16 g9 - 16 g
CalciumCalcium 0.1 - 0.3 g0.1 - 0.3 g
Inorganic phosphateInorganic phosphate 1 -1.5 g1 -1.5 g
AmmoniaAmmonia 0.3 –0.3 – 1 g1 g
UreaUrea 25 – 30 g25 – 30 g
Uric acidUric acid 0.6 –0.6 – 1 g1 g
CreatineCreatine 60 – 150 mg60 – 150 mg
Ketone bodiesKetone bodies 3 – 15 mg3 – 15 mg
16. UreaUrea
25 - 30 g / day25 - 30 g / day
Chief end product of Protein metabolismChief end product of Protein metabolism
Represents 80 – 90% urinary nitrogenRepresents 80 – 90% urinary nitrogen
Quantity varies with amount of protein intakeQuantity varies with amount of protein intake
Urea formation occurs in liverUrea formation occurs in liver
2 molecules of ammonia and 1 molecule of CO2 is2 molecules of ammonia and 1 molecule of CO2 is
converted to urea in each cycleconverted to urea in each cycle
Increased – fever , starvation , adrenocorticalIncreased – fever , starvation , adrenocortical
hyperactivityhyperactivity
Decreased – in liver dysfunction , proteinDecreased – in liver dysfunction , protein
malnutrition and pregnancymalnutrition and pregnancy
17.
18.
19.
20. AmmoniaAmmonia
Fresh urine contains no ammonia – butFresh urine contains no ammonia – but
formed if kept for some timeformed if kept for some time
It is a means to neutralizw ions in the urineIt is a means to neutralizw ions in the urine
Secretes from distal part of renal tubule inSecretes from distal part of renal tubule in
acidosisacidosis
But not in renal acidosis where it fallsBut not in renal acidosis where it falls
Starvation and diabetes – excess secrettionStarvation and diabetes – excess secrettion
21.
22.
23. Creatine and CreatinineCreatine and Creatinine
Creatine is present in Muscle ,Brain and BloodCreatine is present in Muscle ,Brain and Blood
In free and in form of Creatine phosphateIn free and in form of Creatine phosphate
Creatinine is an anhydride of creatineCreatinine is an anhydride of creatine
It is the end productIt is the end product
It is filtered and secreted by the tubulesIt is filtered and secreted by the tubules
Decreases – renal and post renal diseasesDecreases – renal and post renal diseases
JAFF’s reaction – leads to the production ofJAFF’s reaction – leads to the production of
yellowish red colour with an alkaline picrateyellowish red colour with an alkaline picrate
reagentreagent
24. Uric acidUric acid
End product of Purine metabolismEnd product of Purine metabolism
Adenine and guanineAdenine and guanine
Purine rich diet + breakdown of nucleoproteinsPurine rich diet + breakdown of nucleoproteins
A constantly high Uric acid –Gout , LeukemiaA constantly high Uric acid –Gout , Leukemia
Imp in the determination of uric acid calculiImp in the determination of uric acid calculi
Azotemia – high Plasma non-Protein NitrogenAzotemia – high Plasma non-Protein Nitrogen
1.1. Increased tissue protein catabolismIncreased tissue protein catabolism
2.2. Increased break down of blood proteinIncreased break down of blood protein
3.3. Decreased excretion of UreaDecreased excretion of Urea
25. Routine urine examinationRoutine urine examination
Normal Urine does not contain detectableNormal Urine does not contain detectable
amounts of Bilirubin and Glucoseamounts of Bilirubin and Glucose
Single Random sample – qualitative examination –Single Random sample – qualitative examination –
to be examined 1 – 2 hrsto be examined 1 – 2 hrs
Cyclic albuminuria – examine various samples atCyclic albuminuria – examine various samples at
various intervals during day 24various intervals during day 24
24 hr sample24 hr sample
First morning Sample – sample of choice – it isFirst morning Sample – sample of choice – it is
more concentratedmore concentrated
After noon sample – UrobilinogenAfter noon sample – Urobilinogen
Mid stream – bacteriological cultureMid stream – bacteriological culture
26.
27.
28. Discarding the urineDiscarding the urine
If suspected Pathogenic organismIf suspected Pathogenic organism
Add twice the amount of 5% Phenol and letAdd twice the amount of 5% Phenol and let
stand for 2 hoursstand for 2 hours
32. Physical examination of UrinePhysical examination of Urine
Volume = Night : Day = 1:2 to 1: 4Volume = Night : Day = 1:2 to 1: 4
Polyuria >2500 mlPolyuria >2500 ml
1.1. DMDM
2.2. DIDI
3.3. Excessive fluid intakeExcessive fluid intake
4.4. DiureticsDiuretics
5.5. Absorption of exudatesAbsorption of exudates
6.6. Absorption of edema fluidsAbsorption of edema fluids
7.7. HydronephrosisHydronephrosis
33. AnuriaAnuria
1.1. Renal collapseRenal collapse
2.2. Severe Acute nephritisSevere Acute nephritis
3.3. BurnsBurns
4.4. Transfusion reactionTransfusion reaction
5.5. Traumatic shockTraumatic shock
Oliguria <500 mlOliguria <500 ml
1.1. Decreased fluid intakeDecreased fluid intake
2.2. Fluid loss during haemorrhage , vomiting , diarrhoeaFluid loss during haemorrhage , vomiting , diarrhoea
3.3. During the formation of exudatesDuring the formation of exudates
4.4. EdemaEdema
5.5. Acute nephritisAcute nephritis
Nocturia - Early sign of renal impairmentNocturia - Early sign of renal impairment
34. ColourColour
Pale yellow – normal - urochromePale yellow – normal - urochrome
Straw colour Urine is normal –low specificStraw colour Urine is normal –low specific
gravity,<1.010gravity,<1.010
Straw colour fluid – Abnormal – 4+ sugar –Straw colour fluid – Abnormal – 4+ sugar –
urine is like water—high specific gravityurine is like water—high specific gravity
Amber colour urine – normal – due to highAmber colour urine – normal – due to high
specific gravity > 1.020…and out put <1 litrespecific gravity > 1.020…and out put <1 litre
per dayper day
35. Pale colour – DM , DI , Chronic Renal FailurePale colour – DM , DI , Chronic Renal Failure
Orange – fever , bile pigment , medicines likeOrange – fever , bile pigment , medicines like
SennaSenna
Reddish – Hematuria , HaemoglobinuriaReddish – Hematuria , Haemoglobinuria
,Myoglobinuria ,Porphyria,Myoglobinuria ,Porphyria
Brownish Black – Alkaptonuria , MelanoticBrownish Black – Alkaptonuria , Melanotic
tumors ,Poisoning with Lead and mercurytumors ,Poisoning with Lead and mercury
Milky White – Chyluria ,prfesnce of fat globuleMilky White – Chyluria ,prfesnce of fat globule
Green – Bile pigment ,Riboflavin intakeGreen – Bile pigment ,Riboflavin intake
,Methylene Blue,Methylene Blue
Smoky Brown – blood pigmentSmoky Brown – blood pigment
36. Colour of the normal urine darkens onColour of the normal urine darkens on
standingstanding
Beets will turn the urine redBeets will turn the urine red
Rhubarb can cause the urine to turn brownRhubarb can cause the urine to turn brown
Pyridium ,AmidopyrinePyridium ,Amidopyrine orangeorange
PhenylhydrazinePhenylhydrazine dark browndark brown
Triamterine –potassium-sparing diureticTriamterine –potassium-sparing diuretic
Bright yellowBright yellow
Methylene blue ,Amytriptyline – Pink toMethylene blue ,Amytriptyline – Pink to
brownbrown
Iron salts – dark colourIron salts – dark colour
37.
38. AppearanceAppearance
Alkaline urine may be cloudy due toAlkaline urine may be cloudy due to
presence of phophatespresence of phophates
Acidic urine may be cloudy – due toAcidic urine may be cloudy – due to
presence of uratespresence of urates
Presence of WBC ,RBC ,epithelial cellsPresence of WBC ,RBC ,epithelial cells
Presence of mucus , Fat and ChylePresence of mucus , Fat and Chyle
39. OdourOdour
Faintly aromatic –volatile organic acidsFaintly aromatic –volatile organic acids
Fruity – DKAFruity – DKA
Putrid –UTIPutrid –UTI
Fecal odour – Ecoli cystitisFecal odour – Ecoli cystitis
Musty – phenylketonuriaMusty – phenylketonuria
15 cm away – swing from one side to other15 cm away – swing from one side to other
side of the noseside of the nose
40. PHPH
AcidicAcidic
1.1. Metabolic acidosisMetabolic acidosis
2.2. Respiratory AcidosisRespiratory Acidosis
3.3. High Protein intake and ingestion of acidic fruitsHigh Protein intake and ingestion of acidic fruits
4.4. E coli infectionE coli infection
AlkalineAlkaline
1.1. Respiratory Alkalosis ,Metabolic alkalosis ,Respiratory Alkalosis ,Metabolic alkalosis ,
2.2. UTI due to Proteus and PseudomonasUTI due to Proteus and Pseudomonas
Urine should be kept alkaline while treating withUrine should be kept alkaline while treating with
SulphonamidesSulphonamides
This is to prevent the formation of urates andThis is to prevent the formation of urates and
calcium oxalate crystalscalcium oxalate crystals
Proteins will not get precipitated in Alkaline UrineProteins will not get precipitated in Alkaline Urine
41. Always drop of urine into the litmus paperAlways drop of urine into the litmus paper
Specific gravitySpecific gravity
Measure the concentrating and diluting capacityMeasure the concentrating and diluting capacity
of Kidneyof Kidney
1.003 – 1.0301.003 – 1.030
Specimen – if ordered separately – the patientSpecimen – if ordered separately – the patient
should fast for 12 hrsshould fast for 12 hrs
HypersthenuriaHypersthenuria
1.1. DMDM
2.2. DehydrationDehydration
3.3. EclampsiaEclampsia
4.4. ProteinuriaProteinuria
5.5. Lipoid NephrosisLipoid Nephrosis
42. Hyposthenuria < 1.007Hyposthenuria < 1.007
1.1. PyelonephritisPyelonephritis
2.2. HypertensionHypertension
3.3. DIDI
4.4. Protein malnutritionProtein malnutrition
5.5. DiureticsDiuretics
Isosthenuria – chronic renal failure – 1.010Isosthenuria – chronic renal failure – 1.010
SG is highest in first morning sampleSG is highest in first morning sample
Drugs – inc SG – Dextran & Radio opaqueDrugs – inc SG – Dextran & Radio opaque
Contrast mediaContrast media
SG inversely prop to TemperatureSG inversely prop to Temperature
43. SG estimationSG estimation
1.1. Urinometer methodUrinometer method
2.2. Refractometer methodRefractometer method
3.3. Dip – stick methodDip – stick method
Urinometer methodUrinometer method
Principle of buoyancyPrinciple of buoyancy
Inc solute concentrationInc solute concentration increased upthrustincreased upthrust
1.000 – 1.060 with divisions of 0.001 – 0.0021.000 – 1.060 with divisions of 0.001 – 0.002
25 ml capacoty25 ml capacoty
Leave 5 cm at topLeave 5 cm at top
Remove froth from top using filter paperRemove froth from top using filter paper
Take the reading from the lowest meniscusTake the reading from the lowest meniscus
Store –in fresh de – ionized water in the cylinderStore –in fresh de – ionized water in the cylinder
44.
45. CorrectionCorrection
Nl temp – 20CNl temp – 20C
+0.001 for every 3C rise+0.001 for every 3C rise
Albumin -0.003/g%Albumin -0.003/g%
Glucose -0.004 / g%Glucose -0.004 / g%
Correction for dilutionCorrection for dilution
Multiply the last two numbers of the recordedMultiply the last two numbers of the recorded
specific gravity by the dilution factorspecific gravity by the dilution factor
Dis adv – require large volume of urineDis adv – require large volume of urine
Turbid urine makes the reading difficultTurbid urine makes the reading difficult
47. Normal protein – 30 – 150 mg / 24 hoursNormal protein – 30 – 150 mg / 24 hours
Low molecular weight proteins are filterd atLow molecular weight proteins are filterd at
glomerulus - < 60 000glomerulus - < 60 000
Prevents Albumin 69 000Prevents Albumin 69 000
Prevents Gamma globulin 180 000Prevents Gamma globulin 180 000
Normally Protein is not detectable by chemicalNormally Protein is not detectable by chemical
methodsmethods
Renal Tubule secretes a mucoprotein – TammRenal Tubule secretes a mucoprotein – Tamm
Horsfall Protein – normally excreted in urineHorsfall Protein – normally excreted in urine
ProteinuriaProteinuria
1.1. Glomerular DamageGlomerular Damage
2.2. Impaired reabsorption process in tubulesImpaired reabsorption process in tubules
Test for ProteinsTest for Proteins
48. CystinosisCystinosis is ais a lysosomallysosomal storage diseasestorage disease
characterized by the abnormal accumulation of thecharacterized by the abnormal accumulation of the
amino acidamino acid cystinecystine[1][1] . It is a. It is a genetic disordergenetic disorder thatthat
typically follows antypically follows an autosomalautosomal recessiverecessive
inheritanceinheritance pattern. Cystinosis is the mostpattern. Cystinosis is the most
common cause ofcommon cause of FanconiFanconi syndromesyndrome in thein the
pediatric age group. Fanconi syndrome occurspediatric age group. Fanconi syndrome occurs
when the function of cells inwhen the function of cells in renal tubulesrenal tubules areare
impaired, leading to abnormal amounts ofimpaired, leading to abnormal amounts of
carbohydratescarbohydrates andand amino acidsamino acids in thein the urineurine,,
excessive urination, and low blood levels ofexcessive urination, and low blood levels of
potassiumpotassium andand phosphatesphosphates..
49. Lysosomal cystine appears to amplify andLysosomal cystine appears to amplify and
alteralter apoptosisapoptosis in such a way that cells diein such a way that cells die
inappropriately, leading to loss of renalinappropriately, leading to loss of renal
epithelial cells. This results in renal Fanconiepithelial cells. This results in renal Fanconi
syndrome[3] , and similar loss in othersyndrome[3] , and similar loss in other
tissues can account for the short stature,tissues can account for the short stature,
retinopathy, and other features of theretinopathy, and other features of the
disease.disease.
50. There are three distinct types of cystinosis each withThere are three distinct types of cystinosis each with
slightly different symptoms: nephropathic cystinosis,slightly different symptoms: nephropathic cystinosis,
intermediate cystinosis, and non-nephropathic orintermediate cystinosis, and non-nephropathic or
ocular cystinosis. Infants affected by nephropathicocular cystinosis. Infants affected by nephropathic
cystinosis initially exhibit poor growth and particularcystinosis initially exhibit poor growth and particular
kidney problems (sometimes called renal Fanconikidney problems (sometimes called renal Fanconi
syndrome). The kidney problems lead to the loss ofsyndrome). The kidney problems lead to the loss of
important minerals, salts, fluids, and other nutrients.important minerals, salts, fluids, and other nutrients.
The loss of nutrients not only impairs growth, butThe loss of nutrients not only impairs growth, but
may result in soft, bowed bones (hypophosphatemicmay result in soft, bowed bones (hypophosphatemic
rickets), especially in the legs. The nutrientrickets), especially in the legs. The nutrient
imbalances in the body lead to increased urination,imbalances in the body lead to increased urination,
thirst, dehydration, and abnormally acidic bloodthirst, dehydration, and abnormally acidic blood
(acidosis).(acidosis).
51. By about age two, cystine crystals may alsoBy about age two, cystine crystals may also
be present in the cornea. The buildup ofbe present in the cornea. The buildup of
these crystals in the eye causes anthese crystals in the eye causes an
increased sensitivity to light (photophobia).increased sensitivity to light (photophobia).
Without treatment, children with cystinosisWithout treatment, children with cystinosis
are likely to experience complete kidneyare likely to experience complete kidney
failure by about age ten. Other signs andfailure by about age ten. Other signs and
symptoms that may occur in untreatedsymptoms that may occur in untreated
patients include muscle deterioration,patients include muscle deterioration,
blindness, inability to swallow, diabetes, andblindness, inability to swallow, diabetes, and
thyroid and nervous system problems.thyroid and nervous system problems.
52. The signs and symptoms of intermediate cystinosis are the same asThe signs and symptoms of intermediate cystinosis are the same as
nephropathic cystinosis, but they occur at a later age. Intermediatenephropathic cystinosis, but they occur at a later age. Intermediate
cystinosis typically begins to affect individuals around age twelve tocystinosis typically begins to affect individuals around age twelve to
fifteen. Malfunctioning kidneys and corneal crystals are the main initialfifteen. Malfunctioning kidneys and corneal crystals are the main initial
features of this disorder. If intermediate cystinosis is left untreated,features of this disorder. If intermediate cystinosis is left untreated,
complete kidney failure will occur, but usually not until the late teens tocomplete kidney failure will occur, but usually not until the late teens to
mid twenties.mid twenties.
People with non-nephropathic or ocular cystinosis do not usuallyPeople with non-nephropathic or ocular cystinosis do not usually
experience growth impairment or kidney malfunction. The onlyexperience growth impairment or kidney malfunction. The only
symptom is photophobia due to cystine crystals in the cornea.symptom is photophobia due to cystine crystals in the cornea.
It is currently being researched at UC San Diego, The University ofIt is currently being researched at UC San Diego, The University of
Michigan, Tulane University School of Medicine, and at the NationalMichigan, Tulane University School of Medicine, and at the National
Institutes of Health in Bethesda, Maryland as well as at Robert GordonInstitutes of Health in Bethesda, Maryland as well as at Robert Gordon
University in Aberdeen and in Sunderland, UK as well as the NeckerUniversity in Aberdeen and in Sunderland, UK as well as the Necker
Hospital in Paris.Hospital in Paris.
53.
54. Test for proteinTest for protein
All based on precipitation or coagulation byAll based on precipitation or coagulation by
heatheat
Urine should be acidic – if alkaline addUrine should be acidic – if alkaline add
Glacial acetic acid drop by drop – other wiseGlacial acetic acid drop by drop – other wise
coagulation of proteins will not occurcoagulation of proteins will not occur
If SG is very low add sodium Chloride –If SG is very low add sodium Chloride –
other wise it can cause precipitation ofother wise it can cause precipitation of
mucoproteinsmucoproteins
55. Tests for proteinTests for protein
1.1. Heat and Acetic acid testHeat and Acetic acid test
2.2. Sulphosalicylic acid testSulphosalicylic acid test
3.3. Heller’s Nitric acid testHeller’s Nitric acid test
4.4. Dip stickDip stick
5.5. Test with Esbach’s rwagentTest with Esbach’s rwagent
6.6. BiuretBiuret
56. Heat and Acetic acid testHeat and Acetic acid test
Proteins are denatured and coagulated byProteins are denatured and coagulated by
heatheat
1- 2drops of 3% acetic acid1- 2drops of 3% acetic acid
If turbidity due to phosphates or carbonate –If turbidity due to phosphates or carbonate –
ppt disappear after adding Glacial aceticppt disappear after adding Glacial acetic
acidacid
If muco protein – add Nitric acidIf muco protein – add Nitric acid
When tube heated ppt occurs –disappearsWhen tube heated ppt occurs –disappears
when BP is reachedwhen BP is reached BJPBJP
57.
58. Bence Jones ProteinBence Jones Protein
Consisits of Kappa or Lamda light chainsConsisits of Kappa or Lamda light chains
Molecular weight 44 000 – easily filteredMolecular weight 44 000 – easily filtered
Henry Bence Jones first detected it in 1847Henry Bence Jones first detected it in 1847
MMMM
Ppts at 40 -60CPpts at 40 -60C
Dissolves at 100CDissolves at 100C
Reappear at coolingReappear at cooling
Mucin will give a false positive resultMucin will give a false positive result
Heated above 70CHeated above 70C filtered outfiltered out repeat the testrepeat the test
59. Tests for BJPTests for BJP
1.1. Heat Coagulation testHeat Coagulation test
2.2. Toluene Sulphonic Acid testToluene Sulphonic Acid test
3.3. Hydrochloric Acid TestHydrochloric Acid Test
60. BJP conditionsBJP conditions
1.1. 40% cases of MM40% cases of MM
2.2. Some tumor metastasisSome tumor metastasis
3.3. CMLCML
4.4. OsteomalaciaOsteomalacia
5.5. Osteogenic SarcomaOsteogenic Sarcoma
6.6. AmyloidosisAmyloidosis
7.7. Walderstrom’s macroglobinaemiaWalderstrom’s macroglobinaemia
8.8. HypertensionHypertension
61.
62. Esbach’s MethodEsbach’s Method
Quantitative examination of AlbuminQuantitative examination of Albumin
Selective proteinuria –Low molecular weightSelective proteinuria –Low molecular weight
proteins like Albumin and transferrin isproteins like Albumin and transferrin is
selectively excretedselectively excreted
Albumin -66000Albumin -66000
Transferrin 76000Transferrin 76000
1.1. MCGNMCGN
2.2. MGNMGN
3.3. FSGNFSGN
4.4. Nephrotic SyndromeNephrotic Syndrome
63.
64. Globulin detectionGlobulin detection
Urine + ammoniumsulphate + liquorUrine + ammoniumsulphate + liquor
ammonia(if acidic)ammonia(if acidic)
Ppted –apparent on standing or filtrationPpted –apparent on standing or filtration
Fibrinogen detection – detection by theFibrinogen detection – detection by the
production of Fibrin clot and observationproduction of Fibrin clot and observation
under microscopeunder microscope
It can also be done by electrophoresisIt can also be done by electrophoresis
65. Test for glucoseTest for glucose
NL – glucose is not detected by chemicalNL – glucose is not detected by chemical
methodmethod
Renal threshold for Glucose – 160 – 180 mg / dLRenal threshold for Glucose – 160 – 180 mg / dL
Causes of HyperglycemiaCauses of Hyperglycemia
1.1. DMDM
2.2. HyperthyroidismHyperthyroidism
3.3. MIMI
4.4. Cerebral haemorrhageCerebral haemorrhage
5.5. Brain tumorsBrain tumors
6.6. Disease of pancreasDisease of pancreas
66. Renal glycosuriaRenal glycosuria
Defective reabsorption ability of the renalDefective reabsorption ability of the renal
tubuletubule
Lowered renal thresholdLowered renal threshold
Also Occur inAlso Occur in
1.1. CystinosisCystinosis
2.2. Heavy metal poisoningHeavy metal poisoning
3.3. Fanconi’s syndromeFanconi’s syndrome
67. Non pathogenicNon pathogenic
Transient hyperglycemiaTransient hyperglycemia
1.1. PregnancyPregnancy
2.2. Stress and AnxietyStress and Anxiety
3.3. Alimentary GlycosuriaAlimentary Glycosuria
69. Benedict’ testBenedict’ test
Glucose containing an aldehyde group reducesGlucose containing an aldehyde group reduces
Alkaline Cupric Sulphate at higher temperatureAlkaline Cupric Sulphate at higher temperature
to the red cuprous oxideto the red cuprous oxide
1.1. Copper sulphateCopper sulphate
2.2. Sodium carbonateSodium carbonate
3.3. Sodium citrate – prevents the precipitation ofSodium citrate – prevents the precipitation of
cupric hydroxide / cupric carbonatecupric hydroxide / cupric carbonate
4.4. WaterWater
Ketone bodies must be tested if 3+ or 4+Ketone bodies must be tested if 3+ or 4+
70. ProcedureProcedure
5 ml BR5 ml BR
Add 8 drops of urineAdd 8 drops of urine
Heat in water bath – 5 – 10 minHeat in water bath – 5 – 10 min
/ under burner for 2-3 min/ under burner for 2-3 min
Cool under the tap waterCool under the tap water
74. DMDM
Type 1 – Coxsackie B and MumpsType 1 – Coxsackie B and Mumps
HLA DR4HLA DR4
HLA B 8HLA B 8
HLA DR 3HLA DR 3
C peptide Protein assay – more sensitiveC peptide Protein assay – more sensitive
than insulin assay –its level is not affectedthan insulin assay –its level is not affected
by insulin therapyby insulin therapy
77. Principle – Sodium notropruside isPrinciple – Sodium notropruside is
decomposed in alkaline mediumdecomposed in alkaline medium
oxidising substancesoxidising substances forms complxes withforms complxes with
liq.ammonia ..in presence of acetic acid andliq.ammonia ..in presence of acetic acid and
aceto acetic acidaceto acetic acid purple ringpurple ring
Some amount of crystals should remainSome amount of crystals should remain
undissolvedundissolved
Urine is saturated in order to keep liquorUrine is saturated in order to keep liquor
ammonia above the urine solutionammonia above the urine solution
Salicylic acid – false positive - do it beforeSalicylic acid – false positive - do it before
and after heating the urine – nly it should beand after heating the urine – nly it should be
–ve after heating(acetone)–ve after heating(acetone)
78. Bile pigment ,Bile salt ,urobilinogenBile pigment ,Bile salt ,urobilinogen
Unconjugated Brn is water insoluble – cannot be filteredUnconjugated Brn is water insoluble – cannot be filtered
thru glomerulithru glomeruli
Normally very small concentration of conjugated Brn isNormally very small concentration of conjugated Brn is
present in blood – 0.2 -0.4 g/dLpresent in blood – 0.2 -0.4 g/dL
10 – 15% of UBN is reabsorbed into blood stream10 – 15% of UBN is reabsorbed into blood stream
Nl excretion rate of UBN =1-4mg in 24 hoursNl excretion rate of UBN =1-4mg in 24 hours
Freshly collected normal urine gives a positive test forFreshly collected normal urine gives a positive test for
urobilinogenurobilinogen
Nl level of serum bilirubin = 1 mg / dLNl level of serum bilirubin = 1 mg / dL
0.5 mg direct , 0.5 mg indirect0.5 mg direct , 0.5 mg indirect
Total Brn > 2.5 g/dlTotal Brn > 2.5 g/dl icterusicterus
79. Test for BilirubinTest for Bilirubin
Fouchet’s testFouchet’s test
Urine containing Brn – beer brown colourUrine containing Brn – beer brown colour
Yellow foam when shakenYellow foam when shaken
On standing BrnOn standing Brn Bvn …turns greenBvn …turns green
Test for Brn will not be +ve in presnce ofTest for Brn will not be +ve in presnce of
BvnBvn
Hence the specimen should be examinedHence the specimen should be examined
fresh and should be protected from lightfresh and should be protected from light
81. PrinciplePrinciple
BaCl +sulphateBaCl +sulphate BaSO4BaSO4
BS adsorbs the bilirubinBS adsorbs the bilirubin
FR – Ferric Chloride + Trichloro acetic acidFR – Ferric Chloride + Trichloro acetic acid
Ferric chloride oxidises BrnFerric chloride oxidises Brn BvnBvn green colourgreen colour
Certain substances readt with ferric chloride –Certain substances readt with ferric chloride –
salicylates ,urobilin , indican , urobilinogensalicylates ,urobilin , indican , urobilinogen
If urine is acidicIf urine is acidic add few drops of acetic acidadd few drops of acetic acid
Iodine ring test – specific test but not sensitive testIodine ring test – specific test but not sensitive test
False positive –ChlorpromazineFalse positive –Chlorpromazine
False negative – ascorbic acid and nitrateFalse negative – ascorbic acid and nitrate
82. Bile salts – hays testBile salts – hays test
Bile salts reduce the surface tension ofBile salts reduce the surface tension of
urineurine
False positve – Chloroform ,turpentineFalse positve – Chloroform ,turpentine
,thymol,thymol
83. Test for UrobilinogenTest for Urobilinogen
Fresh sampleFresh sample
Ehrlich’s test – 10 ml urine + 1 ml of EREhrlich’s test – 10 ml urine + 1 ml of ER
Para dimethyl amino benzaldehyde + HClPara dimethyl amino benzaldehyde + HCl
UBN + PdMAb – in acidic medium to form a pinkUBN + PdMAb – in acidic medium to form a pink
colourcolour
False positive – Porphobilinogen –False positive – Porphobilinogen –
Watson and Schwartz test – specific forWatson and Schwartz test – specific for
porphobilinogensporphobilinogens
Fresh urineFresh urine to room temtto room temt otherwise Indoxylotherwise Indoxyl
(warm aldehyde )(warm aldehyde ) false +vefalse +ve
84. Test for occult bloodTest for occult blood
Blood benzidine test – determines the freeBlood benzidine test – determines the free
haemoglobin from lysed RBChaemoglobin from lysed RBC
If RBC nopt lysed test –veIf RBC nopt lysed test –ve
+ve hematuria ,haemoglobinuria+ve hematuria ,haemoglobinuria
,myoglobinuriaurine,myoglobinuriaurine
Urine that has low SG <1.007 or highlyUrine that has low SG <1.007 or highly
alkaline can cause lysis of RBCalkaline can cause lysis of RBC
85. HematuriaHematuria
1.1. Acute infectionsAcute infections
2.2. Chronic GNChronic GN
3.3. Nephrotic SyndromeNephrotic Syndrome
4.4. Toxic damage to glomerulusToxic damage to glomerulus
5.5. InfarctionInfarction
6.6. Renal calculiRenal calculi
7.7. Trauma to kidneyTrauma to kidney
8.8. Acute cystitisAcute cystitis
86. MyoglobinuriaMyoglobinuria
1.1. MIMI
2.2. Infarction of large skeletal musclesInfarction of large skeletal muscles
3.3. Destruction of muscle due to crush injuey ,Destruction of muscle due to crush injuey ,
heat stroke ,electric shockheat stroke ,electric shock
4.4. Trauma including beating ,polymiositis andTrauma including beating ,polymiositis and
convulsionsconvulsions
87. Benzidine testBenzidine test
2 ml urine – boil and cool2 ml urine – boil and cool
Another tube –take a pinch of benzidine +3 mlAnother tube –take a pinch of benzidine +3 ml
glacial acetic acidglacial acetic acid
Add 1 ml hydrogen peroxideAdd 1 ml hydrogen peroxide
Mix allMix all
Transinte greenish blue colourTransinte greenish blue colour
Boil and cool – to destroy the peroxidase action ofBoil and cool – to destroy the peroxidase action of
pus cells and bacteriapus cells and bacteria
False –ve = ascorbic acid – an oxygen acceptorFalse –ve = ascorbic acid – an oxygen acceptor
88. Peroxidase activity of HaemoglobinPeroxidase activity of Haemoglobin H2o2H2o2
--. Nascent oxygen--. Nascent oxygen converts Benzidine toconverts Benzidine to
green blue colored complexgreen blue colored complex
90. GalactoseGalactose
By hydrolysis of lactoseBy hydrolysis of lactose
1.1. Mucic acid testMucic acid test
2.2. Phloroglucin testPhloroglucin test
3.3. Orthotoludine testOrthotoludine test
94. Porphobilinogen and porphyrinogens arePorphobilinogen and porphyrinogens are
colourless substancescolourless substances
Oxidised forms are colouredOxidised forms are coloured
Urine with large amount of porphyrine – portUrine with large amount of porphyrine – port
wine colourwine colour
95. 5 hydroxy Indole Acetic Acid5 hydroxy Indole Acetic Acid
Metabolit of serotoninMetabolit of serotonin
Nl – 1- 5 mg in 24 hoursNl – 1- 5 mg in 24 hours
Increased in carcinoid syndromeIncreased in carcinoid syndrome
350 mg / 24 hours350 mg / 24 hours
