Microscopic examination of Urine sediments.ppt

Microscopic Examination Of Urine sediments in the laboratory
prepared by Sintayehu Ambachew
at University of Gondar and University of Adelaide

1. CHAPTER FIVE
Microscopic Examination Of Urine
112/22/2022 By Sintayehu Ambachew Wondemagegn

2. Chapter Objective
At the end of this chapter the students will be able to:
Describe microscopic examination for urine sediment
Describe normal and abnormal urine sediments with their
diagnostic features
Describe formation and significance of casts
Normal and abnormal crystals encounter in urine
sediments
Describe relationship between sediments, chemical,
physical findings in urine
Describe reporting of urinary sediments
Describe quality control in urinalysis
212/22/2022 By Sintayehu Ambachew

3. Chapter Outline
Microscopic Examination of Urine
•Materials&Procedureformicroscopicexamination
•UrinarySediments
OrganizedUrinarySediments
Non-OrganizedUrinarySediments
•Methodsofreportingformedelements
•Sourceoferrorsinthemicroscopicexaminationof
urine
•Qualitycontrolinurinalysis
312/22/2022 By Sintayehu Ambachew

4. Microscope
412/22/2022 By Sintayehu Ambachew

5. Introduction
•Microscopic examination of the urinary sediment
–Most time-consuming part of the routine urinalysis
–To detect and identify insoluble materials present in the
urine
These include RBCs, WBCs, epithelial cells, casts,
bacteria, yeast, parasites, mucus, spermatozoa, crystals,
and artifacts
–Some of these components are of no clinical significance
and others are considered normal unless they are
present in increased amounts
–Both identification and quantitation of the elements must
present
512/22/2022 By Sintayehu Ambachew

6. CONT’….
Valuableinformationcanbeobtainedfrom
microscopicexaminationofurinespecimen
Iftheurinesampleisproperlycollected
Ifthetestiscarefullyperformed,and
Ifthepersonperformingthetestis
knowledgeable,skillful
612/22/2022 By Sintayehu Ambachew

7. Standardization
•Aspectsofmicroscopicexaminationthatshouldbe
standardized:
1.Volumeofurineanalyzed
2.Lengthandforceofcentrifugal
3.Re-suspendingvolumeandconcentrationof
sediment
4.Volumeandamountofsedimentexamined
5.Terminologyandreportingformat
712/22/2022 By Sintayehu Ambachew

8. Principles of method
•Centrifuge10-12mlurinespecimenat1500–2000rpm
for3-5minutes
–Discardthesupernatantbyquickinversionofthetube,
suspend
–TakethesedimentbyPasteurpipettefromthetube
–Transferadropintothecleananddryslide
–Finallyexamineunderthemicroscope
•Repeatedevaluationofurinesedimentisfrequently
valuableinfollowingthecourseandmanagementof
urinarytractdisorders
•Theappearanceofcellularelements,andcastsinthe
urineisareflectionofchangesthattakeplaceinthe
kidney
812/22/2022 By Sintayehu Ambachew

9. Reagents and equipment
•Assembleallnecessarymaterialsusedforthecollection,
centrifugationandexamination
–Specimencollectingcup
–Centrifuge
–Conicalcentrifugetubes
–Pasturepipettewithrubberfitorautomaticpipettes
ifpossible
–Slidesandcoverslides22x22mm.
–Microscope
–Somestainingreagentsifneeded(10%KOHand
crystalviolet,safraninstain,etc)
912/22/2022 By Sintayehu Ambachew

10. Cont’…
1012/22/2022 By Sintayehu Ambachew

11. Specimen requirement
•Preparation of patient
–Instruct the patient how to collect the specimen
–Explain the purpose of the test by using simple language
Do not use medical terms or try to explain details of the
procedure
–If the patient is female, advice her to wash her genital organ
before giving the specimen
This is because bacteria that are normally found on the genital
tract may contaminate the sample and affect the result
•The first morning urine or mid-stream urine specimen is
more preferable, because it is more concentrated and
formed elements are less likely disintegrated
1112/22/2022 By Sintayehu Ambachew

12. Cont’ .…
•Thecollectedurinesampleshouldarriveatadiagnostic
laboratoryassoonaspossible
•Iftheurinesampleisdelayedbymorethan2hours,without
preservation,urinesedimentappearanceandconstituentmay
bechangedandfalseresultsmaybeobtainedandreported
•Ifitisdifficulttodeliverwithin2hrs,itisbettertopreserve
specimenintherefrigeratoratthetemperaturebetween2-6
0
C
orusechemicalpreservatives
Whatarethechangesthatoccurinoldurinespecimen?
1212/22/2022 By Sintayehu Ambachew

13. 1312/22/2022 By Sintayehu Ambachew

14. Source of Errors
•Dryingofthespecimenontheslide
•Ifthesupernatantfluidaftercentrifugationisnotpoured
offproperly
•Ifthewholesedimentwithsupernatantisdiscarded
duringinvertingdownthetubeforlongperiod,the
wholesedimentswillbediscardedandsoagainfalse
negativeresultwillbereported
–Thusanothersampleshouldbecollectedandthetest
shouldberepeated
1412/22/2022 By Sintayehu Ambachew

15. Classification of Urinary Sediments
•Urine sediments can grossly be categorized into 2
Based on the substances they are composed
1.Organized
2.Non-organized sediments
1512/22/2022 By Sintayehu Ambachew

16. Organized (Formed ) elements
RBCs/HPF
WBCs/HPF
Epithelial cells / LPF
Casts / LPF
Parasites/LPF
T. vaginalis/HPF
Bacteria / HPF
Yeast Cells /HPF
Spermatozoa
Miscellaneous substances
1612/22/2022 By Sintayehu Ambachew

17. Non-organized (Non-living Material)
•Slightly acidic urine
•Acidic, Neutral, or slightly alkaline Urine crystal
•Alkaline, Neutral, or Slightly acidic urine
–Triple phosphates
–Amorphous Urates
–Calcium carbonate
–Calcium phosphate
–Calcium Oxalate crystals
1712/22/2022 By Sintayehu Ambachew

18. Organized Urinary Sediments: RBCs
•RBCs not usually present in normal urine
•Appearance:
–RBCs:-Smooth, non-nucleated, biconcave disks
–Measure 7-8 m
–Normally RBCs appear in the fresh sample as intact, small and
faint yellowish discs, darker at the edges
•In concentrated urine may be crenatedand became small (5-6
m)
•In diluted urine, RBCs may be turbid and increase in size (9-10
m)
•In alkaline urine, they may be small or entirely destroyed forming
massive of brownish granules
•In diluted and alkaline urine, the red cell will rupture and release
the hemoglobin, leaving faint colorless cell membrane, and are
known as“ghost” cells (shadow of original cells)
1812/22/2022 By Sintayehu Ambachew

19. 19
•This field contains mostly
RBC’s, (hpf)
•Notice many of them show
biconcavity
•Some show variability in
shape
•Some times it is easy to
mistake fungi for RBC’s
•To get rid of RBC’s so that
WBC’s are more visible –
acetic acid is very helpful,
Why?
12/22/2022 By Sintayehu Ambachew

20. Clinical significance of RBCs
WhenthenumberofRBCsisfoundmorethantheirnormalrange,
usuallygreaterthan5RBCs/HPFitmayindicate:
•Presenceofdiseaseconditionsintheurinarytract,suchas:
–Acuteandchronicglomerulonephritis
–Tumorthaterodeanypartoftheurinarytract
–Renalstone
–Cystitis(Inflammationofbladder)
–Prostitis
–Traumaofthekidney
–Traumaticcatheterization
2012/22/2022 By Sintayehu Ambachew

21. Substances confusing with RBCs
Yeast cells, leukocytes, and bubbles may confuse with RBCs
Differentiate by:-
Yeast cells:
–Smaller and are oval in shape flattened
–Vary considerably in size in one specimen
–Have budding at the surface
–Upon addition of 2-5% acetic acid the RBCs will disappear
Bubbles (oil droplets)
–Vary considerably in size,
–Are extremely refractive or shiny
Leukocytes
–Larger and have granular appearance
–Upon addition of 2-5% acetic acid the RBCs will disappear
2112/22/2022 By Sintayehu Ambachew

22. 22
•These can easily be mistaken
for RBC’s
•They are budding yeast,
notice the almost cactus like
appearance of those in the box
•They will not rupture in
acetic acid, RBC’s will
•These may truly be from the
bladder or they may be a
contamination
Yeast cells
cactus
12/22/2022 By Sintayehu Ambachew

23. Interfering factors
•Factors that may result falsely in high number of RBCs,
i.ewithout the presence of actual renal or other normal
physiological disturbances included:
–Menstrual bleeding
–Vaginal bleeding
–Trauma to perianal area in female patients
–Following traumatic catheterization
•Some drugs:
Aspirin ingestion or over dose
Anticoagulant therapy over dose
2312/22/2022 By Sintayehu Ambachew

24. 24
and Leukocytes
12/22/2022 By Sintayehu Ambachew

25. LEUKOCYTES (WBCs)
•Normalrange:0-4WBC/HPF
•Appearance:normally,cleargranulardiscshaped
–Measure10-15m,thenucleimaybevisible
•Inalkalineurine,theymayincreasetheirsizeandbecome
irregular
•Predominantly,polymorphnuclearneutrophilsareseen
•Sometimesbecauseofpredominanceofneutrophilsandthe
occurrenceofbacterialcelltogetherwithpolymorphonuclear
cells,WBCsarecalledpuscells
•WBCs(puscells)maybeseeninclumps
2512/22/2022 By Sintayehu Ambachew

26. Cont’…..
•Neutrophils exposed to hypotonic urine absorb
water and swell
2612/22/2022 By Sintayehu Ambachew

27. Microscopic Exam
•White blood cells
A few are normal
High numbers indicate
inflammationor infection
somewhere along the
urinary or genital tract
27
40x objective
12/22/2022 By Sintayehu Ambachew

28. Clinical significance of WBC
•Increasednumberofurineleukocyteareseenincaseof:
–Urinarytractinfectionsuchasrenaltuberculosis
–Allrenaldisease
–Bladdertumor
–Cystitis
–Prostatis
•Temporarilyincreasednumberofleukocytesarealso
seenduring:
–Fever
–Afterstrenuousexercise
2812/22/2022 By Sintayehu Ambachew

29. How to report the result of WBCs
•Afterobservingthedistributionofleukocytesunder40x
objective,atleast10fieldsofmicroscope
•When0-5leukocytes/HPFareseen--normal
•5-10WBCs/HPFareseen--fewleukocytes/HPF
•10-20WBCs/HPFareseen--->moderateWBCs/HPF
•20-30WBCs/HPFareseen---->manyWBCs/HPF
•Above30WBCs/HPFareseenfullofWBCs/HPF
29
12/22/2022 By Sintayehu Ambachew

30. 3012/22/2022 By Sintayehu Ambachew

31. Epithelial cells
•It is not unusual to find epithelial cells in the urine
•Unless they are present in large numbers or in
abnormal forms, they represent normal sloughing
of old cells
•Three types of epithelial cells are seen in urine:
1.Squamous
2.Transitional (urothelial), and
3.Renal tubular
3112/22/2022 By Sintayehu Ambachew

32. EPITHELIAL CELLS
Thosecomingfromrenalcells:RTEcells
–Sizeissmallascomparedtootherepithelialcells
–Itmeasures10to18minlength,i.e.,slightlylargerthan
leukocytes
–Verygranular
–Haverefractiveandclearlyvisiblenucleus
–Usuallyseeninassociationwithproteinsorcasts
3212/22/2022 By Sintayehu Ambachew

33. EPITHELIAL CELLS …
Cellsfromrenalpelvisandurethraofthekidney
•Sizeislargerthanrenalepithelia’s
•Thosefrompelvisareaaregranularwithsortoftail,
whilethosefromurethraareovalinshape
•Mostofthetimeurethralepitheliaisseenwith
leukocytes
3312/22/2022 By Sintayehu Ambachew

34. EPITHELIAL CELLS…
Bladdercells
–Aresquamousepithelialcells
–Verylargeinsize
–Shapeseemsrectangularandoftenwithirregular
border
–Havesinglenucleus
3412/22/2022 By Sintayehu Ambachew

35. 35
•These are 2 Squamous
Epithelial cellsshown on
high power (hp)
•They are usually large,
flat, colorless cells
•However, there can be
some granularityto the
cytoplasm & the edges
may be rolled
•The nucleusis usually
distinct & centered
12/22/2022 By Sintayehu Ambachew

36. Microscopic Exam
•Epithelialcellsarelarge
andflat
•Normalcellsthatline
theurinaryandgenital
tractorrenaltubules
36
40x objective
12/22/2022 By Sintayehu Ambachew

37. 37
•ThedetailsoftheseTransitional
Epithelialcells(3)aresomewhat
obscuredbythelargenumberof
bacteriapresent
•Theyareusuallyroundwitha
largeroundnucleus
•Note:Theyaresmallerthanthe
SquamousEpithelialcell
12/22/2022 By Sintayehu Ambachew

38. Clinical significance
•Presenceofepithelialcellsinlargenumber,mostly
renaltypesmayindicate:
–Acutetubulardamage
–Acuteglomerulonephritis
–Silicateoverdose
Note:-Thepresenceoflargenumberofepithelialcells
withlargenumberofLeukocytesmayindicateurinary
tractinfections(UTI)
3812/22/2022 By Sintayehu Ambachew

39. Reporting of epithelial cells
•Epithelial cells distribution reported after looking under
10x objective of the microscope.
•Usually they are reported semi quantitatively by saying
–1-3 epithelial cells /LPF
–3-5 epithelial / LPF
–6-14 epithelial / LPF
–15-25 epithelial/ LPF
–Full of epithelial cells / LPF when the whole field of
10 x objective covered by epithelial cells
3912/22/2022 By Sintayehu Ambachew

40. 4012/22/2022 By Sintayehu Ambachew

41. Casts
•The only elements unique to the kidney
•Long cylindricalstructures
•Result from the solidification of material within the lumen
of the kidney tubules
•Formed by precipitation of proteins, and aggregation of
cells within the renal tubules
•Most urinary casts are formed either in the distal
convoluted tubules or in the collecting ducts
•But rare conditions, casts may be found in the proximal
convoluted tubules
4112/22/2022 By Sintayehu Ambachew

42. Cont’….
•Casts formed in the collecting tubules tends to be:
–Very broad
–Usually indicates the significant reduction in the
functional capacity of the nephron and indicate
severe renal damage
•Most of casts dissociate in alkaline urine, and diluted
urine (specific gravity 1.010) even in the presence of
proteinuria
•Most of them are transparent
•Detected using lower power magnification
4212/22/2022 By Sintayehu Ambachew

43. •Condition that favor formation of casts are:
Proteinuria：albumin, mucoprotein (Tamm-
Horsfall protein) (THP)
High salt concentration
Low PH
Low flow rate
Cont’…
4312/22/2022 By Sintayehu Ambachew

44. Cast Composition and Formation
1.Aggregation of Tamm-Horsfallprotein into individual protein
fibrils attached to the RTE cells
2.Interweaving of protein fibrils to form a loose fibrillar
network
3.Further protein fibril interweaving to form a solid structure
4.Possible attachment of urinary constituents to the solid
matrix
5.Detachment of protein fibrils from the epithelial cells
6.Excretion of the cast
•Any elements present in the tubular filtrate, including cells,
bacteria, granules, pigments, and crystals, may become
embedded in or attached to the cast matrix. The types of
casts found in the sediment represent different clinical
conditions 4412/22/2022 By Sintayehu Ambachew

45. Cont’...…
•Major casts types:
–Hyalinecasts
–Epithelialcasts
–White blood cell, and Red blood cell casts
–Granular (coarse and fine)
–Waxes casts
–Fatty casts
4512/22/2022 By Sintayehu Ambachew

46. Cont’……
•Castsinurinarysedimentisanimportantaidinthe
differentialdiagnosisofrenaldisease
•PureHyalinecastsmaybeseeninproteinuriafroma
varietyofcauses
•SmallHyalinecastseentransientlymayoccurwithmarked
exerciseorfebrileconditions
4612/22/2022 By Sintayehu Ambachew

47. Hyaline Casts
•Allhyalinecasthaveaprecipitatedproteinmatrix,so
therehastoberenalproteinuriaforthesetobeformed
•TheproteinuriaispredominatelyTamm-Horsfall
mucoprotein
•LowpH&increasedelectrolyteconcentrationreadily
precipitateTamm-Horsfallprotein
4712/22/2022 By Sintayehu Ambachew

48. Hyaline Casts
48
Hyaline cast in urine as seen with the 40 objective.
12/22/2022 By Sintayehu Ambachew

49. Hyaline Casts
•ChangesinPHandosmolality
•Arising from normal renal tubular secretion of mucoproteins
•Basicmatrixofallcasts
•Mildrenaldisease
•Themostfrequentcastsandclearcylinders
•Difficulttovisualizebybright-fieldmicroscopy
4912/22/2022 By Sintayehu Ambachew

50. Hyaline Casts
50 12/22/2022 By Sintayehu Ambachew

51. Clinical Implication
•Presence of large number of hyaline casts may show
possible damage of glomerular capillary membrane
•This damage permits leakage of protein through
glomerulus and result in precipitate and gel formation
(i.e. hyaline casts) in the tubule
•Thus this may indicate:
–Nephritis
–Meningitis
–Chronic renal disease
–Congenital heart failure
–Diabetic nephropathy
5112/22/2022 By Sintayehu Ambachew

52. Granular cast
•Moresimilarinappearancewithhyalinecastsandin
whichhomogenous,coursegranulesareseen
•Moredense(opaque)thanhyalinecast
•Shorterandbroaderthanhyalinecasts
•Mayrepresentthefirststageofepithelialcellcast
degeneration
•Basedontheamountandtypeofgranules,dividedinto:
Fine(whichmayappeargreyorpaleyellowincolor)
Coursegranularcasts(whichmayappearasdarker)
5212/22/2022 By Sintayehu Ambachew

53. Granular Casts
•Ifcellularcastsstaywithinthetubulesofthenephrondueto
prolongedstasis,thecellswillbegintodegenerate
•ThesecastsarereferredtoasFinelyGranularorCoarsely
GranularCasts
•Usuallytheoriginaltypeofcellentrappedcannotbe
determinedunlessthecellswereRBC&hgbremains
5312/22/2022 By Sintayehu Ambachew

54. 5412/22/2022 By Sintayehu Ambachew

55. •Granular castunder
bright light microscopy
•If this were on your
scope you would want
to reduce the amount
of light by closing the
substagecondenser,
this really shows the
importance of your
lighting!
5512/22/2022 By Sintayehu Ambachew

56. •Mixed Cellular
Granular Cast,
high power
•Notice that the
cells are
degenerating
•This would tend
to be a Course
Granular
5612/22/2022 By Sintayehu Ambachew

57. •Notice the
coarse
granularity is
very noticeable
5712/22/2022 By Sintayehu Ambachew

58. •Coarse
Granular Cast
next to 2
Epithelial
Cells, high
power
5812/22/2022 By Sintayehu Ambachew

59. Clinical significance
•Granular casts may be seen in:
Acute tubular necrosis
Advanced glomerulonephritis
Pyelonephrites
Malignant nephrosis
Chronic lead poisoning
In healthy individuals these casts may be seen after
strenuous exercise
5912/22/2022 By Sintayehu Ambachew

60. Cellular & Other Cast
•Astheproteinconcentratesinthedistaltubule&becomes
stickier,cellscanbecometrapped
•ThesebecomeHyalineCastswithInclusions&whiletheformal
namewouldbeforexampleHyaline-WBCCast,theyare
frequentlysimplyreferredtoasWBCCast
6012/22/2022 By Sintayehu Ambachew

61. White blood cell casts
•Formedbyaggregatesofwhitebloodcellsthattrapped
inproteinmatrixintherenaltubularlumen
•Anexcessofwhitebloodcells,singlyorinclamps,inthe
urinemayindicateinflammation
•Whitebloodcellcastsdefinitelyarerenalorigin
•Theycharacteristicallyseeninacutepyelonephritisand
occasionallyinglomerulonephritis
6112/22/2022 By Sintayehu Ambachew

62. White blood cell casts
•Upper urinary tract Infection
•Inflammation of the kidney
•Pyelonephritis
•Interstitial nephritis
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63. 6312/22/2022 By Sintayehu Ambachew

64. •WBC cast,
high power
field
•Some of the
nuclear lobes
can be seen
6412/22/2022 By Sintayehu Ambachew

65. Red blood cell casts
•Normalrange:-normallynotseeninnormalindividual
•Usually,theyarefoundinhematuria
•Appearbrowntoalmostcolorless
•Usuallydiagnosticofglomerulardiseases
•Formedusuallyafteraccumulationofcellularelement
intherenaltubules
6512/22/2022 By Sintayehu Ambachew

66. Red blood cell casts…
•Indicateglomerulonephritis
•PrimaryglomerulardiseasewithRBC’spassingthedamaged
glomeruliinlargequantities
•Lupusnephritis
•Rapidlyprogressiveglomerulonephritis
6612/22/2022 By Sintayehu Ambachew

67. •RBC cast, high
power
•Notice the cell
membranes are
clearly visible, but
there does not
appear to be a
nucleus
•Notice how difficult
the mucous
threadsare to see,
this might be
improved by
reducing the light a
bit
6712/22/2022 By Sintayehu Ambachew

68. 6812/22/2022 By Sintayehu Ambachew

69. Epithelial Casts
•EpithelialCastsarecomposedlargelyoftubular
epithelialcelldesquamatedwithinthetubule
•Theyoftenappearastworowsifcellsinproteincast
matrix
•Inflammationofthekidneymaycausegreatersloughing
ofrenalepithelialcells,solargenumberofepithelial
castsisindicativeofrenalparenchymaldiseasewith
tubulardamage
6912/22/2022 By Sintayehu Ambachew

70. 7012/22/2022 By Sintayehu Ambachew

71. •Renal Tubular Cast, high
power
•Can be difficult to
differentiate from other
cellular casts & at times
the decision must be
made on other cells in
the sample
•Notice in this cast the
cells have large nuclei
7112/22/2022 By Sintayehu Ambachew

72. Waxy Casts (Renal Failure Casts)
Not seen in normal individuals
Appearance
•Shorter and broader than hyaline casts
•Composed of homogeneous, yellowish materials
•Broad waxy casts
Are from two to six times the width of ordinary waxycasts
Appear waxy and granular
Have high refractive index
May occur from cells (WBC, RBC, or Epithelial) casts, hyaline
casts
7212/22/2022 By Sintayehu Ambachew

73. •This is a Waxy
Cast, on high
power
•Notice the
crackin the
side of the cast,
which is
frequently seen
in Waxy Casts
7312/22/2022 By Sintayehu Ambachew

74. 7412/22/2022 By Sintayehu Ambachew

75. Waxy Casts
Clinicalsignificance
•Waxy casts are found in:
Chronic renal disease
Tubular inflammation and degeneration
Localized nephron obstruction
Malignant hypertension, in diabetic diseases(nephropathy)
•The presence of waxy casts indicates severity of renal
disease
7512/22/2022 By Sintayehu Ambachew

76. Fatty Casts
•Normally not seen in health individuals
Appearance:
•These are casts, which contain fat dropletsinside them
•Fat droplets are formed after accumulation of fat in the
tubular vessels, especially tubular epithelial and finally
disintegrated
•Clinical Implication:
•The occurrence of fat droplets, oval fat bodies, or fat casts
is:
Very important sign of nephritic syndrome
Chronic renal disease
Inflammation and degeneration of renal tubules
lupus and toxic renal poisoning
7612/22/2022 By Sintayehu Ambachew

77. •This is a cast
containing ‘fat’
bodies, high
power
•On wet mount
the droplets are
highly refractile
[they bounce the
light back]
7712/22/2022 By Sintayehu Ambachew

78. Reporting of casts
•Casts are examined under 10x objective of the
Microscope
•Casts are reported quantitatively by saying:
–Few casts / LPF
–Moderate casts / LPF and
–Many casts / LPF
•During the report the, type of cast that is seen should also
be mentioned
•Example: few hyaline casts / LPF are seen
7812/22/2022 By Sintayehu Ambachew

79. PARASITES
•Parasitesthatcanbeseeninurinemicroscopyare:
Trichomonasvaginalis
Schistosomahaematobium
Wuchereriabancroftie
•OtherparasitessuchasEntrobiousvermiculariesalso
mayoccurduetocontaminationoftheurinewithstool.
7912/22/2022 By Sintayehu Ambachew

80. 80
•Here is another frame of the
Trichomonas(hp), both of
these are shown with phase
microscopywhich enhances
the details of cells with low
refractive indices
12/22/2022 By Sintayehu Ambachew

81. 81
Yeast cells and Trichomonas
vaginalisin urine sediment as
seen with the 40 objective.
12/22/2022 By Sintayehu Ambachew

82. 82
•Trichomonas(hp) is a small
parasite that is very active in
a fresh specimen
•They have multiple flagella
(white pointers) as well as an
undulating membrane which
contribute to movement
•In the absence of movement
they can be mistaken for
other cells
12/22/2022 By Sintayehu Ambachew

83. 83
Schistosoma haematobiumand Enterobiusvermicularis
Egg
12/22/2022 By Sintayehu Ambachew

84. YEAST CELL
•Yeast cells are fungi that are not normally seen in health
individuals.
•Appearance
–Variable in size
–Colorless.
–Oval in shape, and usually form budding.
–Have high refractive index.
–Usually confused with Red Blood Cells.
8412/22/2022 By Sintayehu Ambachew

85. 85
Yeast cells and pseudo hyphae of Candida
albicansin urine sediment as seen with the
40x objective.
12/22/2022 By Sintayehu Ambachew

86. 86
•These can easily be mistaken for
RBC’s
•They are budding yeast, notice the
almost cactus like appearance of
those in the box
•They will not rupture in acetic acid,
RBC’s will
•These may truly be from the bladder
or they may be a contamination
12/22/2022 By Sintayehu Ambachew

87. 87
•These are branching
pseudohyphaeof a
fungus
•Their main significance is
that they obscure more
important features of the
specimen & may indicate
that the specimen is not
a clean catch
12/22/2022 By Sintayehu Ambachew

88. Clinical Significance
•Theyareusuallyofcandidaspecies(candida
albicans)andarecommoninpatientswith
Urinarytractinfection
Vaginitis
Diabeticmellitus
Intensiveantibiotic
immunosuppressivetherapy
8812/22/2022 By Sintayehu Ambachew

89. BACTERIA
•Bacteriaarecommonlyfoundinurinespecimen
becauseofabundantnormalmicrobialfloraofthevagina
orexternalurinarymeatus
•MostcommoncauseofUTIdipsticktestcangive
indirectclue
•Furthertheobservedbacterialcellcanbeidentifiedby
bacteriologicalculture
8912/22/2022 By Sintayehu Ambachew

90. BACTERIA …
•Usuallyonlyasingletypeoforganismispresentin
uncomplicatedacuteurinaryinfections.
•Morethanonetypeoforganismisoftenseeninchronic
andrecurringinfections.
•Vaginalcontaminationofthespecimenisindicatedbya
mixedbacterialflora(includingGrampositiverods)and
oftenthepresenceofepithelialcells.
9012/22/2022 By Sintayehu Ambachew

91. Clinical Significance
•PresenceofbacteriamayindicatethepresenceofUTI
orcontaminationbygenitalorintestinalmicroflora.
•ReportoftheResult
–Fewbacteria/HPF
–Moderatebacteria/HPF
–Manybacteria/HPF
–Fullofbacteria/HPF
9112/22/2022 By Sintayehu Ambachew

92. Bacteria…
•Neisseriagonorrhoeaeinurine
–Inmalepatientswithacuteurethritis,itisoften
possibletomakeapresumptivediagnosisof
gonorrhoeabyfindingGramnegativeintracellular
diplococciinpuscellspassedinurine
9212/22/2022 By Sintayehu Ambachew

93. 93
Crystals in Sediment
12/22/2022 By Sintayehu Ambachew

94. 94
Crystals in Sediment
Microscopic solids usually composed of a small
number of different ions/molecules
Formed by precipitation of urine solutes including
Inorganic salts
Organic compounds, and
Medications (iatrogenic compounds)
Small amount of most type of crystals are not
necessarily pathologic
•High specific gravity specimenscrystals
12/22/2022 By Sintayehu Ambachew

95. 95
Cont’……
•Formation of crystals is most dependent up on:
Concentration of ions and molecules
Urine PH
Decreased flow of urine through tubules
Solutes precipitate more readily at low temperatures
12/22/2022 By Sintayehu Ambachew

96. Urinary crystals identification
•Liver disease
•Inborn errors of metabolism
•Renal damage caused by crystallization of iatrogenic
compounds within the tubules
Crystals are usually reported as rare, few, moderate,
or many per HPF
Abnormal crystals may be averaged and reported per
LPF
9612/22/2022 By Sintayehu Ambachew

97. General Identification Techniques
Characteristic shapes and colors
Urine PH
Polarized microscopy
Solubility characteristics of the crystals
(temperature, chemical)
9712/22/2022 By Sintayehu Ambachew

98. 98
Crystals solubility in urine PH
Solubility of solutes differ at different PH
•Inorganic salts: oxalate, phosphate, calcium,
ammonium & magnesium less soluble in neutral or
alkaline urine
•Organic solutes: uric acid, bilirubin, & cystineless
soluble in acidic urine
•N.B:-Most clinically significant crystal are found in acid
urine
–Include: cystine, tyrosine, leucine& iatrogenic
crytsals: sulfonamide & ampicillin
12/22/2022 By Sintayehu Ambachew

99. Crystals Seen in Acidic Urine
•The most common crystals seen in acidic urine are
urates:
Amorphous urates
Uric acid
Acid urates and
Sodium urates
•Calcium oxalate crystals are frequently seen in acidic
urine, but they can be found in neutral urine and
even rarely in alkaline urine
9912/22/2022 By Sintayehu Ambachew

100. 100
Amorphous Urates
•Amorphous urates
–Non crystalline uratesalts of sodium, potassium,
magnesium & calcium
–Small & yellow-brown granules -similar to sand
–Enhanced by refrigeration
–Can be in acidic or neutral urine(pH greater than 5.5)
12/22/2022 By Sintayehu Ambachew

101. 101
Amorphous Urates
–Will dissolve in alkaline or heated to 60
0
C
–If acetic acid added, uric acid crystals will precipitate
out
–Uroerythrin deposits on uratecrystals giving pink-
organishcolor-referred to as “brick dust”
12/22/2022 By Sintayehu Ambachew

102. Amorphous urates
10212/22/2022 By Sintayehu Ambachew

103. 103
Uric Acid Crystals
•Seen in a variety of shapes, including rhombic,
four-sided flat plates (whetstones), wedges, and
rosettes
•Most common form is diamond shape but may be
cube shaped or cluster in rosettes
•Appear yellow-brown
•Urine pH usually around 5.0 to 5.5
12/22/2022 By Sintayehu Ambachew

104. 104
Uric Acid Crystals
•Diamond shape may cluster in rosettes
•Sometimes 6 sided & must be differentiated from
clinically significant cysteine
•Highly birefringent under polarized light, which aids in
distinguishing them from cystinecrystals
12/22/2022 By Sintayehu Ambachew

105. 105
Uric Acid Crystals and Pathology
•Usually yellow to orange-brown
•Are birefringent under polarizing
light
•Can appear normally BUT
–Seen in large #s in gout &
increased purine metabolism
such as cytotoxic drugs
•E.g. patients with Leukemia who
are receiving chemotherapy
12/22/2022 By Sintayehu Ambachew

106. 106
Acid Urine: Calcium Oxalate Crystals
1.Dihydrateform :-colorless, octahedral envelope or
as two pyramids joined at their bases
2.Monohydrate form:-oval or dumbbell shaped
Rare & can mistake for RBC’s
•Both the dihydrateand monohydrate forms are
birefringent under polarized light
12/22/2022 By Sintayehu Ambachew

107. Acid Urine: Calcium Oxalate Crystals
•Normally associated with foods high in oxalic acid, such as
tomatoes and asparagus, and ascorbic acid
•The most common form of calcium oxalate crystals is the
dehydrate forms
•Clumps of calcium oxalate crystals in fresh urine may be
related to the formation of renal calculi
•Monohydrate crystals are pathologically present in ethylene
glycol poisoning & severe chronic renal disease
10712/22/2022 By Sintayehu Ambachew

108. 10812/22/2022 By Sintayehu Ambachew

109. Crystals Seen in Alkaline Urine
•Phosphates represent the majority of the crystals
seen in alkaline urine
–Amorphous phosphate
–Triple phosphate and
–Calcium phosphate
•Other normal crystals associated with alkaline
urine are calcium carbonate andammonium
biurate
10912/22/2022 By Sintayehu Ambachew

110. Amorphous phosphates
•Alkaline or neutral urine
•Microscopically not distinguishable from amorphous
urates
–Distinguishable on urine PH & solubility
–Precipitate whiterather than pink-orangeof
amorphous urates
–Are soluble in acid & will not dissolve when heated
to 60C
•Fine colorless grains with tendency to obscure other
more significant sediment
•Presence enhanced by refrigeration
11012/22/2022 By Sintayehu Ambachew

111. Amorphous phosphates
11112/22/2022 By Sintayehu Ambachew

112. Triple phosphate
(Ammonium Magnesium Phosphate)
112
•Colorless & in different forms
–most common are 3 & 6 sided
‘coffin lids’
–vary greatly in size
–may also see a ‘fern leaf’ form,
feathery
•See in normal healthy individuals but
are often present in formation of
calculi & are associated with UTI
(urea-splitting bacteria)
12/22/2022 By Sintayehu Ambachew

113. Triple phosphate
Triple Phosphate Crystals
prism
12/22/2022 113By Sintayehu Ambachew

114. 114
Calcium Phosphate
•In 2 forms dicalcium& calcium
•Dicalciumcolorless thin prismsin rosettes or star-
shaped ‘stellar phosphates’
–Tend to have tapered or pointed end & the other
squared off
–Calcium phosphates are irregular granular sheets or
plates often resemble degenerating squamous
epithelial cells
12/22/2022 By Sintayehu Ambachew

115. Calcium phosphate
12/22/2022 115By Sintayehu Ambachew

116. 116
Calcium Carbonate
•Very small granular
crystals
•colorless, Usually
found in pairs
‘dumbbell shape’, or
spherical shapes
•Birefringentwith
polarizing light
•Form gas after the
addition of acetic acid
12/22/2022 By Sintayehu Ambachew

117. 117
Ammonium Biurate
•Yellow brown spheres with
striations
•Can have irregular spicules
‘thorny apple’
•In alkaline or neutral urine
•Not significant unless seen in
fresh urine
•Usually seen in old specimens
•Dissolve in acetic acid or
heating to 60
0
C
12/22/2022 thorny appleBy Sintayehu Ambachew

118. Abnormal Urine Crystals
•Abnormal urine crystals are found in acidic urine or
rarely in neutral urine
Bilirubin
Tyrosine
Leucine
Cystine
Cholesterol
11912/22/2022 By Sintayehu Ambachew

119. 120
Bilirubin
•Appear as fine needles, granules, or plates
–Urine is acidic
–Always yellow-brown
–The bile stains the other components of the
sediment
–Presence of the crystals indicate high concentrations
of bilirubin in the urine
12/22/2022 By Sintayehu Ambachew

120. 121
Bilirubin Crystals: Abnormal State
•If you suspect bilirubin crystals are present, the strip reaction
must confirm the presence of bilirubin
–Otherwise the identification is incorrect
•The presence of the positive bilirubin strip &/or the crystals
indicate a pathologic process -are always considered an
abnormal crystal
•May be seen in liver disease
12/22/2022 By Sintayehu Ambachew

121. 122
Amino Acid Crystals
•Tyrosine
–fine, delicate needles,
colorless or yellow
–frequently in clusters or
sheaves [as in stacks of
wheat]
–seen singly or in small
groups
–in acidic urine
–less soluble than leucine, so
found more often
12/22/2022 By Sintayehu Ambachew

122. 123
Leucine
•Highly refractile yellow to brown
spheres in acid urine.
•Have concentric/radial striations
on their surface
•Can be mistaken for fat globules
[or vice versa]
•But will not stain with fat stains
or appear as maltese crossunder
polarization
•Can be seen in urine containing
tyrosine crystals if use alcohol to
‘precipitate’ Bactrim has similar appearance
check patient history
12/22/2022 By Sintayehu Ambachew

123. 124
Amino Acid Crystals and Pathology
•Amino acid crystals are abnormal & seen in
overflow amino aciduria
–Can be seen in rare cases of liver disease, more
likely to reflect inherited metabolic disorder
–Before reporting should be confirmed by
confirmatory tests such as chromatography
12/22/2022 By Sintayehu Ambachew

124. 125
Cystine: Always Abnormal
•Colorless hexagonal plates
–sides may be uneven
•Crystals appear layered
–tend to clump
–primarily seen in acidic urine
–Must be counted
•Can be confused with uric acid
crystals, must confirm
identification with sodium
cyanide
12/22/2022 By Sintayehu Ambachew

125. 126
Cystine: Always Abnormal
•Clinically significant, seen in congenital cystinosis or
cystinuria
–Deposit out in tubules as calculi/stone causing damage
12/22/2022 By Sintayehu Ambachew

126. 127
Cholesterol
•Clear flat rectangular plates with
notched corners
–in acidic urine
–are soluble in chloroform & in
ether
•Rarely seen
•Presence indicates both ideal
conditions for precipitation &
supersaturating:
•Always seen with positive protein
+ fat droplets, fatty casts or oval
fat bodies
•Seen in nephrotic syndrome &
other renal damage
12/22/2022 By Sintayehu Ambachew

127. 128
Confounding Conditions
•Radiopaque contrast medium[diatrizoatemeglumine] can be
mistaken for cholesterol
–Contrast medium will give abnormally high S.G. >1.040
–Not associated with proteinuria or lipiduria
–Cholesterol crystals found with normal S.G.
•Medications
–Can be excreted in high concentrations, resulting in precipitation
–These crystals are termed ‘iatrogenic’
–Proper identification of drug crystals important in alerting to
potential renal tubular damage
12/22/2022 By Sintayehu Ambachew

128. 129
Ampicillin
•Appear as long thin colorless
prisms or needles
•May aggregate in small clusters or
if refrigerated may form large
clusters
•Appear in acidic urine
•Require large dosage for formation,
so rarely seen
12/22/2022 By Sintayehu Ambachew

129. 130
Sulfonamides
•Highly refractile& birefringent
•In acidic urine,
•Should be confirmed before reporting
•Closely resemble ammonium biuratebut
differentiated on
–pH & solubility
–chemical confirmatory test
•Type varies with form of drug prescribed
•Sulfa drugs have been modified to be
more soluble & so crystals rarely seen
12/22/2022 By Sintayehu Ambachew

130. MISCELLANEOUS
Spermatozoa
•Are small structures consisting of a head and tail,
connected by a short middle piece (neck)
•Easily recognized especially if they are motile
•Frequently seen in the urine of males
•They may see in the urine of females, when the urine
collected after coitus usually not reported, unless the
physician has special interest in it
13112/22/2022 By Sintayehu Ambachew

131. 132
Spermatozoa and occasional pus cell in urine sediment as seen
with the 40 objective
12/22/2022 By Sintayehu Ambachew

132. 133
•There are 2 spermin
this frame
•In a fresh specimen,
they are visible due to
their movement
•In an older urine
specimen, they may be
difficult to visualize
12/22/2022 By Sintayehu Ambachew

133. Mucus threads
•Formedbytheprecipitationofmucoproteinincooledurine
•Havefine,fiberlikeappearance
•Wavyinshapeandtaperedatends
•Ifnotexaminedcarefullymayconfusewithhyalinecasts
13412/22/2022 By Sintayehu Ambachew

134. Contaminates and Artifact Structure
•Muscle fibers
•Vegetable cells
•Structure from slide or cover slide
•Fat droplets (other bubbles)
•Oil droplets
•Pollen greens
•Starch granules
13512/22/2022 By Sintayehu Ambachew

135. Methods for Examining Urine Sediments
•UnstainedUrineSedimentPreparation
–Brightfieldmicroscopy
–PhaseContrasts(PC)microscopy
•StainedPreparation
13612/22/2022 By Sintayehu Ambachew

136. 1.A crystal violet safraninstain (sternheimerand
malbin)
•Useful in the identification of cellular elements
•Staining reaction to crystal –violet safraninstain:
RBC –Purple to dark purple
WBC –Cytoplasm -violet to blue
Nucleus –reddish purple
Glitter cells –blue
137
Stained preparation
12/22/2022 By Sintayehu Ambachew

137. Cont’…..
2.Toluidine blue (0.5%)
A metachromatic stain
Provides enhancement of nuclear detail
Differentiate WBCs and RTE cells
3.Lipid Stains
Oil Red O and Sudan III
Triglycerides and neutral fats stain orange-
red, whereas cholesterol does not stain but
is capable of polarization
13812/22/2022 By Sintayehu Ambachew

138. Cont’….
4.Gram Stain
Differentiate between gram-positive (blue) and
gram negative (red) bacteria
A dried, heat-fixed preparation of the urine
sediment must be used
5.Hansel Stain
Consists of methylene blue and eosin Y
Preferred stain for urinary eosinophils
However, Wright’s stain can also be used
13912/22/2022 By Sintayehu Ambachew

139. Cont’….
6.Prussian Blue Stain
Used and stains the hemosiderin granules a blue
color
7.CytoDiachrome stains
Papanicolaou stain
Detection of malignancies of the lower urinary
tract
14012/22/2022 By Sintayehu Ambachew

140. 14112/22/2022 By Sintayehu Ambachew

141. Automations in Urinalysis
•Automationsareutilizedinurinalysislaboratories
•Thesemachinescanbeappliedforphysical,chemical,and
microscopicalanalysisofurine
•Reflectance photometry
UF-100AutomatedUrineCell
Analyzer
14212/22/2022 By Sintayehu Ambachew

142. Cont’…….
Advantages of automations:
The readings are more reproducible and unbiased
Help to analyze a great number of specimen in less
time
Help to develop standards about the sediments
Give better interpretation about the sediments in
close agreement between laboratories
14312/22/2022 By Sintayehu Ambachew

143. Quality assurance in urinalysis
Quality lab results require
following procedures in
many areas.
Pre-analytic
Analytic
Post-analytic
Quality
Requirements
Standards
12/22/2022 144By Sintayehu Ambachew

144. Definition of Terms
Quality Assurance (QA)–steps taken to assure reliable
laboratory results
Quality control (QC) –procedure, samples and rules to
determine if analysis is acceptable
Pre-analytic: steps before testing the analyte
Analytic: steps of testing the analyte
Post-analytic: steps after testing is complete
12/22/2022
145By Sintayehu Ambachew

145. Quality Assurance?
Assuring quality laboratory service requires Monitoring
and Tracking all aspects:
Pre-analytical
Analytical
Post-analytical
Internal quality control and External quality
assessment
12/22/2022 146By Sintayehu Ambachew

146. Key Components of QA
Internal quality assessment (IQA)
External quality assessment (EQA)
Standardization of processes and procedures (pre-
analytic, analytic and post-analytic phases)
Management and organization
12/22/2022 147By Sintayehu Ambachew

147. 1.Internal Quality Assessment
Monitoring lab procedures
Track lab processes
Instrument calibration
Equipment maintenance
Tracking patient test results
Tracking the reports and archiving
Senior technologist
12/22/2022 148By Sintayehu Ambachew

148. 2.External Quality Assessment
External agency for
1.Proficiency testing
2.On-site evaluation
3.Retesting (rechecking
samples)
12/22/2022 149
On-site evaluation
By Sintayehu Ambachew

149. Quality Assurance
3.Standardization of Processes and
Procedures
Reproducible lab results
Uniform activities
Standard operating procedures
(SOP)
•Pre-analytical
•Analytical
•Post-analytical
12/22/2022 150By Sintayehu Ambachew

150. 151
Summary
•Youshouldbeabletodescribe:
–AppearanceandclinicalsignificanceofRBCandWBC.
–Appearanceandclinicalsignificanceofthreetypesof
epithelialcells.
–Formation,compositionandclinicalsignificanceofthe
differenttypesofurinarycasts.typesofcrystals,
identifythemandstateclinicalsignificanceofeach.
–Otherformedelementstoinclude:bacteria,fat,fibers,
mucous,parasites,sperm,starch,trichomonasand
yeast.
–Typesofqualityassuranceinurinalysis
12/22/2022 By Sintayehu Ambachew

151. Exercise
1.State two technical errors in sediment preparation that
could produce decreased sediment constituents ?
2.The finding of yeast cells in the urine is commonly
associated with____
3.Why do casts vary in size and composition? What is the
primary constituents that all casts have in common?
4.State three factors that contribute to the formation
urinary casts, and explain the significance of each
5.State two methods for enhancing nuclear detail and
two methods for detecting lipids
6.State the advantage of checking complete urinalysis
correlation
12/22/2022 152By Sintayehu Ambachew

152. References:
•Districtlaboratorypracticeintropicalcountries.2
nd
ed.PartI.
MonicaCheesbrough,2005
•Textbookofurinalysisandbodyfluids.DorisLR,AnnEN,1983
•Urinalysisandbodyfluids:Acolortextandatlas.KarenMR,Jean
JL.1995
•Clinicalchemistry:Principles,procedures,correlation.3
rd
ed.
MichaelL.Bishopetal.1996
•TietzTextbookofclinicalchemistry.3
rd
ed.CarlAB,EdwardRA,
1999
•Clinicalchemistry:Theory,analysis,correlation4
th
ed.Lawrence
AK.2003
•Urinalysislecturenote.MistireW.,DawiteY.
•Urinalysisandbodyfluids/SusanKingStrasinger,5thed.2008
15312/22/2022 By Sintayehu Ambachew

153. 15412/22/2022 By Sintayehu Ambachew

154. Case Studies and Clinical Situations
1.An 85-year-old women with diabetes and a broken hip has been confined
to bed for the past 3 months. Results of an ancillary blood glucose test
are 250 mg/dL, and her physician orders additional blood tests and a
routine urinalysis.
The urinalysis report is as follows:
COLOR: Pale yellow, KETONES: Negative
CLARITY: Hazy, BLOOD: Moderate
SP. GRAVITY: 1.020, BILIRUBIN: Negative
pH: 5.5, UROBILINOGEN: Normal
PROTEIN: Trace, NITRITE: Negative
GLUCOSE: 100 mg/dL, LEUKOCYTES: 2
Microscopic:
20 to 25 WBCs/hpf
Many yeast cells and hyphae
12/22/2022 155By Sintayehu Ambachew

155. Cont’….
A.Why are yeast infections common in patients with diabetes
mellitus?
B.With a blood glucose level of 250 mg/dL, should glucose be
present in the urine? Why or why not?
C.Is there a discrepancy between the negative nitrite and the
positive leukocyte esterase results? Explain your answer.
D.What is the major discrepancy between the chemical and
microscopic results?
E.Considering the patient’s history, what is the most probable
cause for the discrepancy?
12/22/2022 156By Sintayehu Ambachew

156. 2.A 2-year-old left unattended in the garage for 5 minutes is
suspected of ingesting antifreeze (ethylene glycol).
The urinalysis has a pH of 6.0 and is negative on the chemical
examination.
Two distinct forms of crystals are observed in the microscopic
examination.
A.What type of crystals would you expect to be present?
B.What are the two crystal forms present?
C.Describe the two forms.
D.Which form would you expect to be predominant?
12/22/2022 157By Sintayehu Ambachew

157. 3.As supervisor of the urinalysis section, you are reviewing results.
State why or why not each of the following results would concern
you.
A.The presence of waxy casts and a negative protein in urine from a 6-
month-old girl
B.Increased transitional epithelial cells in a specimen obtained
following cystoscopy
C.Tyrosine crystals in a specimen with a negative bilirubin test result
D.Cystine crystals in a specimen from a patient diagnosed with gout
E.Cholesterol crystals in urine with a specific gravity greater than 1.040
F.Trichomonas vaginalis in a male urine specimen
G.Amorphous urates and calcium carbonate crystals in a specimen
with a pH of 6.0
12/22/2022 158By Sintayehu Ambachew