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CYTOLOGY101:PRINCIPLES,PRACTICESANDCOMMONDIAGNOSES

OFCYTOLOGYINSMALLANIMALMEDICINE
JENNIFERNEEL,DVM,DACVP(CLINICALPATHOLOGY)
JAIMETARIGO,DVM,DACVP(CLINICALPATHOLOGY)
NCSUCVMVETFORUMCONTINUINGEDUCATION
04/11/12

PREPARING&SCREENINGSAMPLESTOMAXIMIZEYIELDFROMYOURCYTOLOGY

Cytologyoffersalowinvasiverapiddiagnostictestforveterinarypatients.Advantagesofcytology
include:rapidresults,inexpensive(comparedwithasurgicalbiopsy),oftenprovidesadefinitive
diagnosisorcannarrowthelistofdifferentials,andcausesminimalpatientdiscomfort.

OBTAININGYOURSAMPLE

Thegoalofpreparingslidesistohaveacellular,thinpreparationofintactcells.
NOTE:Ifalesioniscystic,itisoftenbeneficialtofirstaspirateoffthefluidportion,andthenaspirate
anysolidportionofthelesionleftafterfluidremoval.Bothportionsshouldbeexamined.

FineNeedleAspirates
Techniquesvary,buttissuesamplesaremostcommonlycollectedusinga20,22or25gaugeneedle.
Ifanyglassdust/gritispresent,drywipetheslidespriortouse.
WewillbeshowingyouavideoofDr.Birkenheuerinactionhesubmitscytologythatisconsistently
cellular,intact,anddiagnostic..

TheAspirationTechnique(typicallythefirstapproach):
o Attacha22gaugeneedletoa6ccor12ccsyringe,breakthesealonthesyringe.
o Inserttheneedleintothetissuepullbackto12ccofnegativepressure.
o Gentlypokewhileredirectingandmaintainingsteadynegativepressure.
o Removethesyringeandneedle,taketheneedleoffandfillsyringeto6ccwithair
o Holdingtheneedle1/81/4awayfromtheslideexpelthesamplewithonereasonably
forcefulpush.
o Holdthesampleslideandlayasecondslideacrossperpendiculartothesampleslide.
o Donotapplypressure,allowgravitytodispersethematerial.
o Keepthehandholdingthesampleslideflexible(likeaspringboard)andinonesmooth
motion,slidingforwardandwithaslightdownwardpressureslidethematerialacross.
TheAspirationTechniquewithPulsatileNegativePressure(forlesionsrefractorytoexfoliation)
o Usethetechniqueabovebutafterentryintothetissueapplyrepeatednegativepressure
inbypullingbackontheplungerinapulsatilefashiontonomorethan~2ccofair.
o Releasetheplungerpriortoremovalfromthetissueandusethetechniqueabovetoexpel
andspreadyoursample.
Nonaspirationtechnique(whenthefirstattemptappearsverybloody):
o Inserta22gaugeneedleintothetissueandgentlypokewhileredirecting.
o Removetheneedle,attacha6ccsyringefilledwith6ccofair.
o Usethetechniqueabovetoexpelandspreadyoursample.

NOTE:lubricantmaterialandultrasoundgelstainabrightmagentacolorandcanobscureyour
sampleandmakediagnosisdifficultminmizingtheseonthesurfaceoftheskinbeforeFNAwillhelp.

ADDITIONALSLIDEPREPARATIONTECHNIQUES
Drawbackandpushforwardorwedgetechnique:Thisissimilartomakingabloodsmear.It
workswellforallliquidspecimensincludingtissueandmassaspiratesiftheyareveryfluidanddo
notcontainvisibleflecksoftissue(inthatcase,useasquashtechnique).
Squashtechnique(slidesandwichtechniquewithpressureapplied):Thisworkswellforsamples
clumpsofmucusorcellsarepresent.Thesampleisplacedononeslideandasecondslideis
placedoverit,gentlepressureisappliedandtheslidesarepulledapartsimilarlytothe
techniquesdescribedabove.
Touchimprints.Touchimprintsofbiopsyspecimenscanprovidepreliminaryfindingsandcan
confirmgoodsamplingofthelesion.Note:touchimprintsofulceratedordraininglesionsmay
containsurfacecontaminationorinfectionmaynotberepresentativeoftheunderlyinglesion;if
thereisadiscretemassthatisulcerated,oftenanaspirateofthemassportionisneeded.The
mostcommonproblemsencounteredwithtouchimprintsareinadequateblottingofthe
specimenandnonexfoliativelesions.
o Blotting:Thecellsofinterestwilloftenonlyadheretotheglassifbloodandserumare
removed.Blotthetissuegentlyonapapertoweluntilthetowelbeginstosticktothetissue.If
multipleslidesarebeingmade,youmanyneedtoreblotoften,especiallyforverybloody
tissuessuchasliverorspleen.
Fordelicateorsmallbiopsyspecimens,youmaynothaveenoughsampleand
gentlytouchingaglassslidetothetissueseveraltimesmaystillbeuseful,
especiallyforbonemarrowsbecausetheytendtoexfoliateverywell,evenwithout
blotting.
Forfirmersamplesthatmaynotexfoliatewellyoucangentlyroughenthesurface
byscrapingascalpelbladeoverit,thenmakingtouchimprints

AREYOURSLIDESWORTHSENDINGOFFFORCYTOLOGY?
1. Dotheyhaveenough(orany)cells?Arethecellsarebroken?Isonlybloodispresent?Isitthe
righttissue?
Examinetheslideonlowpower(4xor10x)Lookforareaswithintactcellsthatarespreadout
adequatelyforevaluationyoucanuseredbloodcellsiftheyrethereasareferenceiftheyre
pileduporcondensedyourecellsofinterestwillbealso.
2. Examineseveralareasathighpower(40or50x).
Ifyouareusinga40xobjective,remembertoplaceadrycoverslipontopofthecytology
specimenbeforeyouviewitthisobjectiveisdesignedtobeusedwithacoverslipandwithout
onethingswilllookblurry.

ALOGISTICALAPPROACHTODIAGNOSINGLYMPHNODEASPIRATES

Thenicethingaboutlymphnodesistherearereallyaverylimitednumberofdiagnosesavailablesoit
iseasytodevelopasystematicapproachtolymphnodeevaluation.Thethreebasicfindingsfor
lymphtissueare1)normalorunremarkable2)reactive/hyperplasticand3)lymphoma.Additional
lymphnodefindingsincludeinflammation,metastaticneoplasiaornondiagnosticspecimens
(acellular,aspirationofperinodalfat,salivarytissue).

Step1:Sizinglymphocytes
Therealkeytodiagnosingnormal,reactiveandneoplasticlymphtissueisbeingabletoaccurately
sizelymphocytes,soyourfirststepisalwaystofindameasuringstickandthentrytoidentifyasmall
lymphocyte.Todothis,youwillfirstneedtofindathinareawithintactcellsdontbotherlooking
inthethickregionstosizeyourlymphocytes,youwontbeaccurate.Therearefewhelpful
measuringsticksthebestmeasuringstickisaneutrophil.Asmalllymphocytewillbesmallerthana
neutrophil,willhavecoarse,darkchromatinandjustatinyamountofcytoplasm,oftenvisibleon
onlyonesideofthecell.Cantfindaneutrophil?Inapinch,youcanuseawellspreadoutredcell.In
dogs,RBCsareapproximatelythesamesizeorslightlysmallerthanasmalllymphocyte(thiswont
holdtrueincatsorotherspecieswhohavesmallerRBCs).Onemoremeasuringstick;thenucleusofa
plasmacellisapproximatelythesizeofasmalllymphocyte.Intermediatesizedlymphocytesare
approximatelyneutrophilsizedtoslightlylarger,theyhavealittlemorecytoplasmandthechromatin
isalittlemoreopen(dispersed,stainspalerandislessclumped).Largelymphocytesareupto4
timesthediameterofaredcellwithpalelacychromatin,athinrimofcytoplasmand,dependingon
thelevelofmaturity,mayormaynothavenucleoli.

NORMALLYMPHNODE
Innormallymphnode,approximately7595%ofcellsaresmall,maturelymphocytes.The
remainderofcellsconsistsprimarilyofintermediatesizedlymphocyteswithlownumbersoflarge
lymphocytes.Rareplasmacellsmaybeseen;nucleiareround,similarinsizetosmalllymphocytes,
areeccentricallylocatedinthecytoplasmandhaveverycoarsechromatin.Thecellshavemoderate
amountsofdeeplybasophiliccytoplasmwithaperinuclearclearzone(golgiapparatus).Mottcells
areplasmacelldistendedwithlargevacuolesofimmunoglobulin(Russellbodies).Raremastcells
maybeseen.Thesearelargeroundtoovalcellswithround,centralnucleiandabundantbasophilic
topurplecytoplasmicgranuleswhichoftenobscurethenucleardetail.Granulesmaynotstainwith
DiffQuik.Youmayseeafewhistiocytes,otherinflammatorycellsshouldbeproportionatetothe
degreeofhemodilutionpresent.Lymphoglandularbodiesarepresentinthebackground.Theseare
plateletsizedbasophilicfragmentsofcytoplasmwhichpinchoffofproliferatinglymphocytes.While
notspecificforlymphocytes,theyarehighlycharacteristicoflymphtissue.

HYPERPLASTIC/REACTIVELYMPHNODE
Lymphoidhyperplasia(proliferationofthelymphoidpopulationsecondarytoantigenicstimulation)
andreactivity(expansionoftheplasmacellpopulation)canbelocalizedorgeneralized.Localized
reactioncanbeduetoanysourceofinflammationdrainingtothatlymphnodeincludingskindisease,

anabscess,injuryorevenaninflamedneoplasm.Generalizedreactionscanoccurwithrickettsial
diseases(RMSF,ehrlichiosis/anaplasmosis),FeLVandFIVoranysystemicinflammatorycondition.

Oneofthefirstcluesthatyouaredealingwithareactive/hyperplasticlymphnodeisyour
observationfromlowpower.Whatyouwillnoticeisthatthereisalotofvariationincellsizeand
chromatintexture/color(darkandcoarselyclumpedtopaleandopen)whereasnormaland
lymphomanodesaremoremonotonousappearingfromlowpower.Specifically,smalllymphocytes
stillpredominatebutintermediatetolargelymphocytesareincreasedandtheremaybeincreased
numberofplasmacells+/Mottcells.Increasesinothercelltypesarevariable;youmaysee
increasednumbersofmacrophagesandmastcells.Neutrophilsandeosinophilsmayalsobe
increasedbutrepresentlessthan5%and3%ofthenucleatedcellcountrespectively(ifgreater,then
nodeisinflamed).Anormalcytologicappearanceinalymphnodethatgrosslyisincreasedinsizecan
alsobeconsistentwithhyperplasia.

Atypicalhyperplasiacanoccur.Inthiscondition,thepredominantcelltypeisintermediateinsize
ratherthansmall,andthiscanbedifficultorimpossibletodistinguishfromsmall/intermediatecell
lymphoma(seebelow).AtypicalhyperplasiaisseenmostoftenindogswithEhrlichiosisbuthasalso
beenreportedinotherdiseasesuchasLeishmaniasis.Atypicalhyperplasiaoftheperipherallymph
nodesinyoungFeLVpositivecatsissimilarandcanlooklikelymphomabecautiousabout
diagnosinglymphomaintheperipherallymphnodeofacat,itisveryuncommon.

LYMPHADENITIS
Thisischaracterizedbyincreasednumbersofneutrophils(>5%),eosinophils(>3%),macrophagesor
MNGC(nosetnumberforthese).Namethelesionbythepredominantinflammatorycomponent.
Reactivehyperplasiaisusuallypresentaswell.Presenceofsuppurativeorpyogranulomatous
inflammationmeansathoroughhuntforetiologicagentsisneededbacteria,protozoa,fungi,algae.
Someneoplasmscaninducesuppurativeinflammationifmetastatictothelymphnode(SCC).

LYMPHOMA
Lymphomaaffectingtheperipherallymphnodesofdogs(iemulticentriclymphoma)isthemost
commonformindogs(80%ofcases).Otherformsoflymphomaexistindogsincluding
craniomediasinal,gastrointestinal,cutaneous,andprimaryextranodalforms.Lymphomaofthe
caninelymphnodecanbeconfidentlydiagnosedwhenlymphoblastsrepresentgreaterthan50%of
thecellpopulation.Inrealitymostcaninelymphomacaseshavesomethingmorelike90%orgreater
blastcells,whichmakesitevenmorestraightforward.Lymphomacanalsooccurintheinternallymph
nodesandorgansofdogs(seebelow).Lymphomaisveryuncommoninfelineperipherallymph
nodesbecautiousaboutdiagnosinglymphomaintheperipherallymphnodeofacat(consultation
withapathologistforthesesamplesisrecommended).

Blastcellsareintermediatetolargesizedroundcellswithlargeroundnuclei,delicatechromatin,
distinctnucleoli.Nucleiaresurroundedbyathinofbasophiliccytoplasmthatmayhavefinevacuoles.
Cellsmayhaveaprominentperinuclearclearzone.Thesecellslookverydistinctivefromlowpower:
Largepalenucleisurroundedbyadarkbluerimofcytoplasm.Keystocorrectlyidentifyingcellsas
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blastsarethefeaturesofimmaturity:visiblenucleolianddispersed/activechromatin(pale,lacey,
delicate).Dontgettoohunguponabsolutesize;evenifthecellisnotmuchbiggerthana
neutrophil,ifithasthesefeatures,itcountsasablast.Normalintermediateorevenslightlylarger
lymphocytesshouldhavecondensingchromatinandshouldnothaveprominentnucleoli.Remember
thatcellsotherthanplasmacellscanhaveaperinuclearclearzone,itonlyindicatesthatthecellis
activelyproducingproteins(plasmacellsareoneofthefewfullydifferentiatedcellswithaprominent
golgiareabecausetheyareveryactivemakingimmunoglobulin).Donotmistakelymphomafora
plasmacelltumorfewplasmacelltumorswillhaveanovertlyblasticappearanceandplasmacell
neoplasmsareinfrequentlyfoundinlymphnodes.Unlikemanymalignantneoplasticpopulations,
lymphomatendstoberelativelyuniforminsize(lessanisocytosisandanisokaryosis).
Lymphoglandularbodiesoftenareincreasedinthebackgroundandmaybelargerandmore
basophilic.Mitoticfigures(normalorabnormal)maybeincreasedinnumber

Therearesomepotentialdifficultiesinthediagnosisoflymphoma.Small/intermediatecell
lymphomasarecharacterizedbysmalltointermediatesizedcellsthatarenotovertlyblastic.

Alimentarylymphomaincatsisoneofthemostcommonexamplesofsmall/intermediatecell
lymphomaandoneofthemostcommonlyoccurringcancersofthecat.Cytologicfindingsofasmall
tointermediatecellpopulationonFNAofintestinesorintestinallymphnodesinthecatdonotrule
outindolentlymphoma.Dr.Suterwillspeaktoyouaboutagreatsolutiontohelpwithlowinvasive
diagnosisusingatestcalledPARR(PCRforantigenicrearrangement)thatcanbeperformedonthe
slidessubmittedforcytology.Otherwise,smalltointermediatecelllymphomasaninfrequently
occurandmaybedifficulttodifferentiatefromhyperplasiabasedoncytology(needhistopathology,
immunocytochemistry,flowcytometricevaluation,receptorgenerearrangementPCR,etc.).

Immunophenotype(BorTcell)cannotbeaccuratelypredictedbasedonmorphologyandprovides
usefulinformationforyouandyourclientasanoftenimportantprognosticfactorindogs.Thiscan
bedeterminedusingmonoclonalantibodiesspecifictoBandTcelllineageonformalinfixedtissues,
unstainedcytologyslidesorviaflowcytometry.LineageandclonalitycanalsobedeterminedbyPCR
evaluationforantigenreceptorgenerearrangement(PARR)fromtissueaspirateorbiopsy,orfrom
cellsscrapedoffofacytologypreparation.HypercalcemiaisclassicallyassociatedwithTcell
lymphomas.MostlymphomasaffectingtheperipherallymphnodesofdogsareBcellorigin.

METASTASTICNEOPLASIA
Thisischaracterizedbythepresenceofaforeigncellpopulation(carcinomacellsforexample)orhigh
numbersofacelltypethatshouldonlybepresentinverylownumbers(mastcellsinthelymph
node).Metastaticcellsmaybepresentinlownumbersanywhereontheslidesoitisvitalthatall
areasofallslidesareexaminedonlowpower;adiagnosticclustercanbepresentanywhere.Ihave
foundthatmetastaticsquamouscellcarcinomacanbeparticularlytricky;itiseasytomissafewsmall
clustersorindividualizedtumorcellsifyouarenotthoroughortryandgotooquickly.Negative
findingsdonotexcludethepossibilityofmetastaticdisease

OTHER
Artifacts.Rupturedcellscanappearasauniformpopulationofintermediatesizedlymphocytes;be
sureyouareevaluatingintactcellsbeforemakinginterpretations.Cellsinthickareastakeupstain
poorlymakingnuclearmorphologydifficulttoassess.Dense/thickregionspreventcellsfrom
spreadingoutmakingsizingdifficult.

PerinodaladiposetissueappearssimilartoalipomaornormalSQfat(seelipomaincutaneouslumps
andbumps).

Salivarytissue.Salivaryglandiscommonlyaspiratedinsteadofthemandibularlymphnode.On
cytology,ithasastreaming,pinkmucinousbackgroundwithprominentwindrowingofRBCs(rowing
upofcells,duetoathickormucinousmaterial,commonlyseeninsalivaryaspiratesandnormaljoint
fluid,namedaftertheappearanceofwindrowsoffieldcropssuchashay).Thisisoftenmucheasier
toappreciateonlowpower.Theglandularepithelialcellsappearasfoamycellswithsmall,round
nucleiandabundantpalecytoplasm.Theyareoftenarrangedinsmallclusters(glandular
epithelium),butbecausethecellsaredelicate,theycaneasilyruptureresultinginnakednuclei
dontmistaketheseforsmalllymphoyctes!Itisimportanttonotmistakesalivarytissuefor
metastaticneoplasia.

COMMONMISTAKESORDIFFICULTIESWITHLYMPHNODES
1. Selectingtoothickofanareaforexamination.Similartoexaminingaperipheralbloodsmear,you
needtochooseanareawherethecellsarespreadoutenoughsoyoucanclearlyseeindividual
cells.Theindividualcellsalsoneedtobespreadoutsometimesthebackgroundinaregionis
thickandthecellscantadequatelyspreadandinsteadappearlikelittledarkbullets;youneedto
beabletoseegoodnuclearandcytoplasmicdetail.Ifcellsareballedupornotspreadout,find
anotherregion.Unlikeaperipheralbloodsmear,thereisnoformulaforfindingthebestareait
canbeanywhereontheslidesoyouneedtospendtimeonlowpowerhuntingforthoseregions.
2. Attemptingtointerpretrupturedcells.Makesureyoucanseeboththenucleusandthe
cytoplasmicbordertobesureyouareevaluatingintactcells.Whencellsrupture,theyswell;the
nucleuswillstainalighterpurpleandwillbesmudgy.
3. Notcriticallyevaluatingcellsize.Toclassifylymphoidpopulations,youneedtobeabletogauge
size,usingthemeasuringsticksmentionedabovewillhelpandeventuallythisbecomessecond
nature.
4. Troubledistinguishingapredominatelyblasticpopulation(lymphoma)fromareactivepopulation.
PaycloseattentiontosizeANDmaturityofthecells(seelymphomasection).Pickgoodareas
blastcellsaredelicateandmaybeeasilydamaged,theyalsostainpoorlywhenintoothickofa
regionmakingidentificationofnucleolidifficult.
5. Differentiatingnucleolifromclumpedchromatin.Clumpedchromatinissimilarincolortotherest
ofthechromatinbutisdarkerandoftenisnotadistinctshapewithasharpmargin.Nucleoliare
slightlytosignificantlydifferentincolor(usuallymorebluevs.chromatinspurplecolor)andarea
distinctshape.Somelymphomashaveverydelicatenucleoli,youneedtopickgoodareasto
examine.
6. Rushingtooil.Onceyougotooil,youhavetostopandcleantheslideinordertogobackto40x
thistypicallydissuadespeoplefromfurtherevaluation.Itsevenworsewithrealcytologypreps
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becauseyoucantsimplywipetheoiloff,youneedtoeithercleantheslideinzyleneorputa
coverslipontopoftheoil.Forthesereasons,oilshouldbethelaststepintheprocess.Utilizethe
10xand40xobjectivestoidentifyandexaminemultipleareasoftheslide.

FLOWCYTOMETRY&IMMUNOPHENTYPING
JonathanE.Fogle,DVM,PhD,DipACVIM

HOWDOESFLOWCYTOMETRYWORK?
Flow cytometry provides a way of analyzing cells based on their size, granularity, and cell surface
markers.Thetablebelowliststheantigenswegenerallystainforandthecellstheyarefoundon.
Usingflowcytometry,wecanimmunophenotypelymphoidcellpopulationstodeterminetherange
ofcelltypeswithinthepopulation.Thediversityofcelltypeswithinthepopulationcanhelp
determineifthecellsareneoplasticorreactive.Apopulationthatexpressesonlyonephenotype
suggestsneoplasiaratherthanareactiveprocess.Aberrantsurfacemarkerexpression(e.g.CD3but
notCD4orCD8,lossofCD45)alsosuggestsneoplasia.Incasesofleukemia,expressionofCD34
suggestsanacute,notachronicdisease.CanineandfelineleukocytestypicallydisplayCD45,whichis
usedinconjunctionwithFSCandSSCcharacteristicstosegregatethelymphocytes,monocytesand
granulocytes.

Whensubmittedforflowcytometry(immunophenotypicanalysis)asampleisdividedintomultiple
tubesforvariouscontrolsandsubjectedtoanalysisusingapanelcomprisedofdifferent
combinationsofantireceptorantibodies(e.g.CD4,CD21etc.).Samplesanalyzedaretypically
reportedaseitherapercentageofpositivecells(dotplots)orasthemeanfluorescenceintensity
(MFI).Theresultsofthisanalysisaretheninterpretedbytheflowcytometrist.Wewillpresent
severalofthemostusefulclinicalexamplesforyouanddiscusstheimmunophenotypingresults.
Itshouldbenotedthatflowcytometryshouldnotbeusedastheonlymethodofdeterminationof
neoplasia. History, clinical signs, and cytology are important and necessary for the laboratory to
correctlyinterprettheflowcytometryresults.Additionally,difficultcasesmayalsoneedanalysisfor
clonalitybyPARR.

ANTIGEN

ANTIGENGENERALLYFOUNDON

CD3
CD4

Tcells
Thelpercells

CD8

Tcytotoxic/suppressorcells

CD21
B5

MatureBcells
SomebutnotallBcells(CDmoleculeunknown)

CD14

monocytes/macrophages

CD34
CD79a/b
CD11d

Hematopoieticprogenetorcells;acuteleukemias
PartoftheBcellreceptor;Abstainsintracellularcomponent;seenonimmature
Bcells
Macrophagesofspleenandbonemarrow;LGLCLLs

CD45

Allleukocytes(lymphsbright;PMNsdim;blastsdimtonegative)

PARRPCRFORANTIGENRECEPTORREARRANGEMENT
StevenSuter,VMD,PhD,DipACVIM(Oncology)

WHATISPARR?
Canine/felinePARRisaPCRbasedassaytodetermineifapopulationofcellsistheresultoftheclonal
expansionofBorTcells,whichusually,butnotalways,implieslymphoidneoplasia.Thetestutilizes
genomic DNA and PCR primers that are specific to the canine V(D)J splice junctions of B and T cell
receptor gene segments in lymphocytes. Because a clonal expansion of a population of neoplastic
lymphocytescanbePARRpositiveforbothBandTcellrearrangements,PARRshouldneverbeused
forthelineageassignmentofcaninelymphoma.Additionally,PARRshouldneverbeusedasthesole
assay to determine if an animal has lymphoma or leukemia. Instead, PARR can be one of the tests
used to interpret difficult cases in addition to cytology, immunocytochemistry, flow cytometry,
history, and clinical signs. The main advantage of PARR is the ability to differentiate a
monoclonal/oligoclonal or neoplastic lymphoid proliferation from a reactive and polyclonal or
pseudoclonalproliferation.ThePARRassaycanberunonfreshlymphnodeaspirates,bonemarrow
aspirates, cellulareffusions, blood, cells scraped from cellular cytology slides, formalin fixedtissues
(25micronsections),andfreshfrozentissues(25micronsectionskeptat80degrees).

WHENISPARRMOSTUSEFUL?

FelinePARR:Anaidinthediagnosisofirritableboweldisorderversuslowgradealimentary
lymphomaincats.
Alimentarylymphomaisoneofthemostcommonlyoccurringcancersofthecatandtheincidence
ofthisdiseasehasincreasedsignificantlyoverthepastdecade.Despitetheprevalenceofthis
disease,appropriatediagnosisandtreatmentcanpresentamajorchallengetoveterinarians.There
aretwomainformsoffelinealimentarylymphomawhichincludethemostcommonsmallcell
lymphocytic,welldifferentiated,lowgradephenotypeandthelargecell,lymphoblastic,highgrade
phenotype.Itcanbedifficulttodifferentiatelowgradelymphomafrominflammatoryboweldisease,
eveninthefaceoffullthicknessbiopsies.Thealgorithmfordistinguishingbetweenalimentary
inflammatoryversusneoplasticdiseaseinthecatoftenincludesabdominalultrasound,fineneedle
aspirates/cytology,endoscopicorfullthicknessbiopsies/histopathology,andimmunophenotyping
(withlymphocytemarkersontissueorflowcytometrysamplesobtainedfromaspirates).Therehas
beenincreasedrecognitionofthevaluetoaddtheFelinePolymeraseChainReactionforAntigenic
Rearrangement(PARR)tothisalgorithm.FelinePARRisaPCRtestforaclonalpopulationofcells
whichissupportiveofaneoplasticpopulation.ThesensitivityoffelineTcellPARRisreportedtobe
~89%.ThisoffersanextremelysensitivemeansofdetectingaclonalpopulationofTcellsthatis
supportiveofalimentarylymphoma.FelineBcellPARRisapproximately60%sensitive.Cats
presentingwithanindolentdiseasecoursewithclinicalsymptomsofirritableboweldisorderarethe
largesttargetpopulationthatPARRmaybenefit.PARRcanbeperformedoncytologysamples
offeringaninitiallowinvasivediagnostictool.PARRcanalsobeperformedonhistopathology
sampleswhenmoreinvasivediagnosticsarerequired.Increasingawarenessaboutwhichcasesthis
testislikelytohelpasadiagnostictool,whichsamplestosubmitandhowtoprocessthemwillaidin
thediagnosisofirritableboweldisorderversuslowgradealimentarylymphomaincats.
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Althoughimmunophenotypeoffelinegastrointestinallymphomavariesintheliteraturearecent
publicationcharacterizingfelinegastrointestinallymphomareportedamajorityofTcellorigin.
(MoorePF,RodriguezBertosA,KassPH.VetPathol.FelineGastrointestinalLymphoma:Mucosal
Architecture,Immunophenotype,andMolecularClonality.VetPathol2011Apr19)89%of103cases
wereofTcellorigin(predominantlysmallcell),11%wereofBcellorigin(predominantlylarge
cell/lymphoblastic)andPARRinthatstudydetected90%oftheTcelllymphomas.

CANINEPARR
Indogs,PARRhasareportedsensitivityandspecificityofapproximately90%.
Usefulapplicationsinclude:
Immunophenotyping*whenflowcytometryisnotaccessibleordefinitive
Suppurtiveevidenceoflymphoidoriginwhenacancerisanaplastic(iedifficulttotell
whothecellreallyis)
DistinguishinghistiocyticdiseasefromLSA
Confirminganeoplasticplasmacellpopulationwhencytologyisnotdefinitive

IMMUNOPHENOTYPEOFCANINELYMPHOMAISIMPORTANT
MostlymphomasaffectingtheperipherallymphnodesofdogsareBcellorigin.Prognosisof
lymphomaindogsdependsonseveralfactors,clinicallythefollowingparametersarecommonly
usedtoassessprognosis:Phenotype.BcellphenotypehasabetterprognosisthanTcell(Bisbad,T
isterrible).Stageofdisease.Thestageoflymphomaindogshasamodestassociationwithprognosis.
StagingisbasedontissuesinvolvedandisgivenaRomannumber(IV).Thestageisfollowedbythe
letteraorbtoindicateabsence(a)orpresence(b)ofclinicalsigns.StageIaisthebest,stageVbisthe
worst.MostdogsarediagnosedatstageIII,IVorV.Stagesareasfollows:
StageI:Involvementislimitedtoasinglenodeorlymphoidtissueinasingleorgan(notbone
marrow)
StageII:Involvementofmanylymphnodesinaregionalarea(withorwithouttonsils)
StageIII:Generalizedlymphnodeinvolvement
StageIV:Liverand/orspleeninvolvement(withorwithoutstageIIIdisease)
StageV:Manifestationintheblood(neoplasticcellsincirculation)andinvolvementofbone
marrowand/orotherorgansystems(withorwithoutstagesI,II,IIIorIVdisease)

Initialresponsetotreatment.Dogsthatrespondpositivelytoinitialinductionhaveabetterprognosis
thanthosewhodonotrespondwell.Manydogswhorespondwellshowsignificantimprovementof
clinicalsignswithin35daysofinduction.ThemediansurvivaltimeforstageIVorVBcelllymphoma
indogsonmultiagentchemotherapyprotocolis1214months,whilethemediatesurvivalfordogs
withstageIVorVTcelllymphomaonmultiagentchemotherapyprotocolis69months.

Prognosisincatsismoredifficultbecauseofthewidevariationintissuesaffected.Overall,compared
withdogs,catsarelesslikelytorespondtotreatment,arelesslikelytohavehighremissionratesand
tendtohaveshortersurvivalperiods.Ingeneral,importantfactorsinprognosisforlymphomaincats
include:1)Responsetotreatmentcatsthathaveacompleteresponsehaveabetterprognosisvs.
thosethathaveapartialresponse,2)FeLVstatuspositiveFeLVstatushasaworseprognosisvs.
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negativestatus,and3)Clinicalstageearlyclinicalstageisbetterthanlateclinicalstage.Stagesareas
follows:
Stage1:Asingleextranodaltumororonenodalarea
Stage2:AsingleextranodaltumorwithregionallymphnodeinvolvementORtwoormorenodal
areasonthesamesideofthediaphragmORtwoextranodaltumorsonthesamesideofthe
diaphragmwithorwithoutregionallymphnodeinvolvementORaresectableprimaryGItumor
withorwithoutassociatedmesentericlymphnodeinvolvement.
Stage3:TwosingleextranodaltumorsonoppositesidesofthediaphragmORtwoormorenodal
areasaboveandbelowthediaphragmORanyprimary,extensive,unresectable,intraabdominal
diseaseORanyparaspinalorepiduraltumor
Stage4:Stage1,2,or3withliverand/orspleeninvolvement
Stage5:Stage1,2,3or4tumorortumorswithinitialinvolvementoftheCNSand/orbone
marrow.

Anatomiclocation.MediastinallymphomainFeLVpositivecatshastheworstprognosis,
only23monthswithchemotherapy.Nasallymphomahasthebestprognosiswithmedian
survivaltimesapproaching1.5yearswithlocalradiationtreatment.Renallymphomahasa
shortersurvivaltimeof36monthswithchemotherapy.Doxorubricin.Additionof
Doxorubricintothechemotherapyprotocolisassociatedwithbettersurvivaltimes.NOTE:
Phenotype(BorT)isNOTaprognosticindicatorinthecat.

FelineGastrointestinalLymphoma120cats(MoorePF,RodriguezBertosA,KassPH.VetPathol.
FelineGastrointestinalLymphoma:MucosalArchitecture,Immunophenotype,andMolecular
Clonality.VetPathol2011Apr19)
o 103(89%)Tcelllymphoma
84mucosal(smallcell)mediansurvival29mos.
19transmuralmediansurvival1.5mos.
9/19=lymphoblastic
alllargelyconfinedtoSI
PARRdetected90%ofTcellcases

o 19(11%)Bcelllymphoma
19transmuralmediansurvival3.5mos.
mostlymphoblastic(acytologicdiagnosis!)
stomach, jejunum, ileoceco-colic junction

CYTOLOGYSLIDESAMPLESSUBMITTEDTONCSUVHCCYTOLOGYSERVICEMAYBESUBMITTEDDIRECTLYFORPARR.
DETAILEDSUBMISSIONINFORMATIONISAVAILBABLEATTHEENDOFTHENOTESSECTIONORAT:
CYTOLOGY: http://www.cvm.ncsu.edu/vhc/csds/Cytology.html
PARR&FLOWCYTOMETRY :http://www.cvm.ncsu.edu/dphp/labs/clinicalimmunologylab.html
GooglingNCSUCytologyorNCSUPARRworkstoo!
11

COMMONLUMPS&BUMPSINSMALLANIMALPRACTICE
Epidermalandfollicularcysts
Thesecystsareverycommon,nonneoplasticlesionslocatedinthedermisofdogsorcatsandmay
representuptoathirdofallnonneoplastictumorlikemassesfromdogs.Theycanbefluctuantor
firm,andaretypicallysmoothandwellcircumscribed.Thedorsumandextremitiesarecommon
locations.Thecystislinedbymaturesquamousepithelialcells;keratinizedepithelialcellsandcellular
debrisaccumulatesinthecenter.Oncytologicexamination,theyconsistofmature,fullykeratinized
squamousepithelialcellsandcellfragments.Maturekeratinizedsquamouscellsareangular,flat,
andbasophiliccellsthathavelosttheirnucleiduringmaturation.Fromlowpower,theseaspirates
havebeendescribedaslookinglikebluecornflakesscatteredontheslide.Thecytoplasmofthe
cellsandcellfragmentsexhibitevidenceofkeratinizationwhichischaracterizedbythick/denseor
waxyappearingcytoplasmandadistinctpaletodeeprobinseggorskybluecolor.Abackground
ofkeratindebrismaybeseen.Cholesterolcrystalsmayalsobepresent.Ruptureortraumawill
induceintensepyogranulomatousinflammationbecausekeratinizedcellsinduceaforeignbody
typeresponse.WheneverIhavepyogranulomatousinflammationfromamasslikelesioninadog,I
alwayslookforclumpsofkeratintodetermineifitlikelyarupturedfollicularorepidermalcyst.
Cytologicallytheselesionsappearsimilartoepithelialtumorswithfolliculardifferentiationthus
bothdifferentialsareapplicable.Theyarealltypicallybenignandexcisioniscurative.

Tumorswithfolliculardifferentiation
Similartoepithelialandfollicularcysts,tumorswithfolliculardifferentiationcanhavelarge
accumulationsofkeratinocytesandkeratindebris.Thereareseveraldifferenttypesoffollicular
tumors(somepeoplecallthemadnexaltumors)withtrichoepitheliomaandpilomatricomabeing
mostcommon.Typically,theyarebenignalthoughmalignantformsdorarelyoccur.Thesetumorsare
usuallyhairless,raised,wellcircumscribedandmaybeulcerated.Tumorsandcystscanhavea
similarcytologicappearanceandmanyclinicalpathologistswillbottomlineallofthemas
epithelialinclusioncyst,butitshouldbeunderstoodthathistopathologyisnecessaryto
distinguishcystsfromtumorsandtoaccuratelyidentifytumortype.Theprimaryfeatureisthesame
asforcysts;thepresenceofmaturesquamousepithelialcellsandcelldebris.
Youmayalsoseeghostcells;keratinizedepithelialcellswithanemptyholeinthecenterwherethe
nucleususedtobe.Thisindicatesaspecialtypeofkeratinizationthatepithelialcellsofthehairbulb
undergoastheyformthehairshaft(matricalkeratinization),sofindinglotsoftheseindicatesyou
eitherhaveafollicularcystoratumorwithfolliculardifferentiation.WhenIseelargenumbersof
ghostcells,Ifavortumor.Ghostcellscanbeindividualizedortheycanpileuptogetherformingcrude
linearstructureslikeanattemptatformingahairbulb,oftentheyhaveasortofhoneycombed
appearancebecauseoftheemptynuclearspaces.Withfolliculartumors,youmayalsoseesmall
amountsofuniformbasilarepithelialtypecellsorsebaceouscellsifthewallofthetumoris
aspirated.
Ruptureorleakageoffolliculartumorsorfollicular/epithelialcystswillproduceintensesuppurative
topyogranulomatousinflammation.Lookforislandsofmaturesquamouscellsamongstthe
inflammatorycells.

12

Basalcelltumor
Thetumorthatforyearsbeencalledbasalcelltumorhasnowbeenreclassified;basalcelltumoras
aspecifictumortypeisnolongerrecognizedindogs(butisstillrecognizedincats).Instead,whatwas
previouslycalledbasalcelltumoroncytologyisnowrecognizedtobeagroupofdifferenttumors
withoverlappingfeaturesincluding,trichoblastomasindogs(tumorofthehairbulb)formerlybasal
celltumorindogs),sweatglandtumors,basalcelltumorsincats(benigntumorofbasalcellsin
cats),andlesscommonlybasalcellcarcinoma(basalcellcarcinomasarelowgradetumorslocally
invasivebutwithlowmetastaticpotential,goodprognosiswithremoval).

Differentiatingthesetumortypestypicallyrequireshistopathology.Thisiswhymostclinical
pathologistwillbottomlineacytologycomposedprimarilyofbasilarappearingcellsascutaneous
basilarneoplasia(orsimilarterminology),butsomewillstillusebasalcelltumorasamoregeneral
term.Theyarealltypicallybenignandexcisioniscurative.

Basalcelltumors(cats)andtrichoblastomas(dogs)arethemostcommonskintumorofcatsandare
alsocommonindogs.Thesetumorsareusuallybenignorverylowgrade.Typically,theypresentasa
solitarydomeshaped,firm,hairlessmass.Theyarefreelymovablebutfirmlyadheredtothe
overlyingskin.Commonsitesindogsandcatsincludethehead,neckandshoulders.

CYTOLOGICALLY:
Thesetumorsareclassicallyepithelialinappearanceandarrangedintightclusterswithalmostno
individualizedcellsseen.Clustersareoftenextremelydenseandverybasophilic.Cellshavea
relativelyuniformappearance,cuboidalshape,andscantamountsofdeeplybasophiliccytoplasm
thatmayoccasionallybepigmented.Somecanhavecysticportions.Aspirationoftheseregions
resultsinabasophilicbackgroundwithscatteredcellulardebris.Oftencholesterolcrystalsormelanin
granulesarepresent,andyoumayormaynothavescatteredclustersofbasilarepithelialcells.Ifyou
findyouhaveaspiratedfluid,youshouldremoveasmuchofthefluidaspossible,thentryaspirating
anyremainingtissuemass.Youmayseesmallregionsofsebaceousorsquamousdifferentiation
individualizedorsmallclustersofsebaceouscellsorfociofkeratinizedepithelialcells.Meibomian
glandadenomaoftheeyemaylooklikeabasalcelltumorwithsebaceousdifferentiation

Difficulty:Othertumorscan,oncytology,appearsimilartobasalcelltumors(i.e.appeartobe
composedofuniformclustersofbasalepithelialcells),soitisimportanttokeepinmindother
potentialtumorsaswell.Locationshouldhelptipyouoffastowhentobemorecautiousiewe
haveseensebaceousepitheliomas(akindofsebaceoustumorwithalotofbasalcells)andsweat
ducttumorswhichbothtypicallybenign)thatlookbasilar.Andwehavealsoseensalivaryductular
carcinomasandmammarytumors(thatcanbemalignant)thatlookbasilarbutlocationshouldtip
youofftobecautious(ieassociatedwithaparticularstructurelikethemammaryglandorsalivary
gland).

Sebaceoushyperplasia
Thisisacommonconditioninolderdogsandcanalsobeseenincats(femalesmorethanmales).It
representsahyperplasticresponseofthenormalsebaceousepithelium.Classiclocationsareonthe
13

headanddorsum.Incats,itscommontofindlesionsatthetailbase.Typicallythegrowthsappearas
araised,hairlesslesionwithacauliflowerappearanceorasanintradermalmultilobulatedmass.
Grossly,thesamplemayappearoilyorgreasywhenunstained.Cytologically,thislesioniscomposed
ofuniformandtightlycohesiveclustersofsebaceousepithelialcells.Cellshavemoderateto
abundant,highlyvacuolated/foamycytoplasmandsmallcentralnuclei.Theremaybesmall
numbersofmorebasalappearingcells(reservecells).Thesearethecellswhicharewaitingto
differentiateintosebaceouscells.Thebackgroundoftheslideisoftenveryvacuolatedfrom
sebaceousmaterialreleasedbyrupturedcells.Cytologyalonecantdifferentiatesebaceous
hyperplasiafromsebaceousadenomathisisgenerallyoflittlesignificanceasadenomashave
similarclinicalfeaturesandabenignbehaviorMeibomianglandadenomaoftheeyemaylooksimilar
cytologically.

Sebaceousadenoma
Behaviorandcytologicappearancearesimilartosebaceoushyperplasia.Cytologyalonecant
differentiatesebaceoushyperplasiafromsebaceousadenomawhichisusuallyoflittlesignificance
asadenomashavesimilarclinicalfeaturestohyperplasiaandusuallyabenignbehavior.
Meibomianglandadenomaoftheeyemaylooksimilarcytologically.Findinghighnumbersofbasilar
typecellsmayindicatethetumorisasebaceousepitheliomaoroneofanumberofothertumors
featuringbasalcellswithregionsofadnexalorsquamousdifferentiation.Thesearealsostilltypically
benignandexcisionwhenneedediscurative.

Lipoma
Lipomasareparticularlycommoninolderdogswhomaydevelopmultipletumors,butcanalsobe
seenincats.Liposarcomasdooccurbutcanbedifficulttodistinguishfromothertypesofsarcomasor
evenneuroendocrinetumors;thesetumorsshouldhaveclearlymalignantfeatures.Lipomaspresent
assofttofirm,wellcircumscribed,freelymovablesubcutaneousmasses;occasionallytheycanbein
deepertissuesandbelessmovable,morefixed.Dogscanalsodevelopinfiltrativelipomas.These
arealsocomposedofwelldifferentiatedadiposetissue,butcaninfiltrateandcausedestructionof
muscleandconnectivetissue.NOTE:othertumorscanbegrosslymistakenforalipoma,Iveseenthis
mostoftenwithMCTandhemangiopericytomas.Grossly,withlipomastheslidesappeargreasyand
donotdry.Theadipocytesappearasclustersofverylargesignetringshapedcellswithabundant
palepinkcytoplasm.Thesecellsareeasytoseeon10xbecauseoftheirsize.Normalsubcutaneous
orperinodaladiposetissuelooksthesame.

AlcoholfixativesusedinRomanowskytypestainswilldissolveawayanyfreelipidandmayclear
awaycellsaswell.Youcanskipthefixativeandstaintheslideswithonlytheredandblueportions
ofquickstainsbutthiscanleadtocontaminationofyourstainsolution.

Squamouscellcarcinoma
Indogs,commonsitesofSCCincludethehead,distalextremities(nailbeds),ventralabdomen,and
perineum.Itsthesecondmostcommonoraltumorofdogs(firstismelanoma).Intheoralcavity,a
rostrallocationhasabetterprognosis(usuallybenign),whereasacaudallocationisgrave,especially
ifsublingualortonsillar.SCCofthenasalplanumisverybad.Incatsitisassociatedwithchronicsolar
14

injuryinanimalswithwhitehair.Commonsitesincludeearpinnae,frontalridges,eyelids,nose,and
lips.Itsthemostcommonoraltumorinthisspeciesandcarriesapoorprognosisinthislocation.

Oftenthesetumorsareulceratedandaccompaniedbysuppurativeinflammation+/asecondary
bacterialinfection.Cellscanbefoundinsheetsor,quitecommonly,asindividualizedcellsorsmall
clustersofcells.Sometimesyouwillseeangularshapedcellsthatarereminiscentofsquamouscells
ofthenormalskin.Youmayfindafewtadpoleshapedcellscellswithatailofcytoplasmgiving
thematadpolelikeshape.Welldifferentiatedtumorswillshowevidenceofkeratinizationdenseor
waxycytoplasmwithacharacteristicrobinseggorskybluecolorofvaryingintensity.
Remember,notallcellswithinatumorwillbenoticeablykeratinized.Fine,perinuclearvacuolization
iscommon.Emperipolesisisoftenseenwhenonecellcrawlsintoanothercellscytoplasm.Hereitis
neutrophilsmigratingintothecytoplasmofneoplasticsquamouscells.ThisisnotspecificforSCC,but
itisoneofthetumortypeswhereImostoftenseeit.

TherearesomedifficultieswithdiagnosingSCC.Cellsofthisparticulartumortypemaybehighly
individualizedandcanbemistakenformesenchymalorroundcelltumors.Thekeyistosearchfor
smallclusterswithdefinitivecellcelljunctionsandrecognizingevidenceofkeratinization.Well
differentiatedtumorswithinflammationcanbedifficulttodistinguishfromdysplasticchanges
inducedinnonneoplasticcellssecondarytochronicinflammationorirritation.Ifyouhavealotof
inflammationwithoutreallystrongcriteriaofmalignancy,becautious.Othertypesoftumorscanalso
keratinize(thyroid,mammary,transitionalcell,papillomas,lungtumors,etc.),sodontjumponSSC
asadiagnosisifitisinanatypicallocationorthepresentationisatypical.

Perianalglandadenoma
Thistumorismostcommoninintact,maledogs(mayactuallybehyperplasiaintheseguysover
90%ofintactdogsarecuredbycastrationplusmassremoval,castrationalonecanresultinpartial
tumorregression)andisrareincats.Itisalsorareinintactfemalesasestrogenstendtosuppress
tumorgrowthwhileandrogenssupportit.Itissometimesreferredtoasahepatoidtumoror
hepatoidcellsbecausetheybeararemarkableresemblancetohepatocytes.Malignanttumorsare
uncommonandusuallyareassociatedwithgreatercellularpleomorphism,althoughwell
differentiated,malignantorinvasivetumorshavebeenreported.

Perianalglandadenomasusuallyappearasslowgrowing(monthstoyears),raised,smoothto
lobulatedmassesneartheanusbutmayalsobefoundanywherealongthemidlineofthedog,
fromnosetotailwiththemostcommonnonperianalregionsbeingtheundersideofthetail,
perineum,prepuse,andcaudalthighs.Theymayulcerateandbecomeinfectediftraumatized.

Thesetumorsaredistinctlyepithelialandarecharacterizedbyverycohesiveclustersofrelatively
uniform,large,ovaltopolygonalcellswithround,somewhateccentricnuclei.Individualizedcells
areuncommon.Theyhaveverycharacteristiccytoplasm:itisabundantandfinely
granular/mottled,pinkandblue.NOTE:overstainingwiththeblueportionofDiffQuikcanresultin
lossofthepinkishgranular/mottledquality.Youmayseeasmallnumbersofreservecells(basilar
typecells)flattenedaroundtheperipheryofthelargehepatoidcellclusters.Thesetumorsarewell
15

vascularized,youwillusuallyhaveabackgroundofblood.Thesearebenigntumorsbutcanbe
difficulttocontrollocallyandexcisecompletelyintheperianal/perinealregionearlyexcisionis
helpful.

Apocrineglandadenocarcinoma.
Thistumorismostcommonlyseeninolderdogsbutisrarelyseenincats.Itisahighlymalignant
tumorthatarisesfromtheapocrineglandsoftheanalsacandtheyoftenmetastasizetoregional
lymphnodesevenwhenthetumorisgrosslysmall(<1cm);anywherebetween4696%havealready
metastasizedatthetimeofclinicaldetection.Grossly,itisafirmsubcutaneoustumorfirmly
attachedtotheanalsac,rectalexammaybenecessarytodetectit,especiallyasmanyhaveinward,
ratherthanoutward,growth.Metastasistotheregionallymphnodes(sublumbaroringuinal)is
common;moredistantmetastasiscanoccurlateindisease.Hypercalcemiaisfoundin2550%of
casesduetoPTHrPproduction.Initialclinicalsigns(polyuriaandpolydypsia,constipation)areoften
relatedtothehypercalcemiaratherthanthetumormass.Monitoringforthereturnofhypercalcemia
aftersurgicalresectioncanbeusedtoscreenforrecurrence.Prognosisisgenerallypoor,butwith
properclinicalmanagementandearlydetection,affectedanimalsmayliveseveralyears.

Althoughconsideredacarcinoma,thistumorhasaclassicallyneuroendocrineappearanceon
cytologylooselycohesiveclustersofcellswithroundnucleiandindistinctcellmargins.Cellsare
easilyrupturedresultinginnumerousnakednucleiinthebackground.Looseacinarorrosettelike
arrangementsmaybeseenwithinclusters.Oftenapocrineglandadenocarcinomascanlook
deceptivelyuniformwithlittleanisocyosisoranisokaryosisseen.Theyhaveroundnucleiwith
delicatechromatinandmayhavesmallindistinctnucleoli.Typicallytheyhaveasmallamountofpale
bluecytoplasmwhichmaybefinelyvacuolated.

Softtissuesarcoma
Sarcomaswillhavefeaturesofmesenchymalcellsandvaryingcriteriaofmalignancy.Sometumors
willaspiratewell,otherwillfollowtherulesformesenchymalcellsandaspiratepoorly.Asofttissue
sarcomaissimplyasarcomaarisingfromthesofttissuesthisisanumbrellatermcoveringavariety
oftumorsthatcanariseinthetissues.Althoughthereareavarietyofdifferentsarcomasthatcanbe
groupedinthesofttissuesarcomacamp,distinguishingbetweenthemisdifficultcytologicallyas
varioustumorscanhavesimilarfeaturessomesofttissuesarcomaswarranthistologicevaluation
tobetterdeterminetumortypeandgrade.Tumorgradeisdirectlycorrelatedwithprognosis.
Gradingisdoneonhistopathologysectionsanddependsheavilyonmitoticindex(moremitotic
figuresisbad).Gradingofsofttissuesarcomasisonascalefrom13.Approximately1015%ofgrade
1and2tumorsand40%ofgrade3tumorswillmetastasize.Grades1and2carryagoodprognosis
withsurgicaland/orradiationtreatment.

Indogs,themostcommonsofttissuesarcomas(inorderoffrequency)arehemangiopericytoma(not
relatedtohemangiosarcomas),fibrosarcomaandperipheralnervesheathtumors,butthereare
manyothertypes.Definitiveidentificationrequiresbiopsyandhistopathology.

16

Incats,themostcommonsofttissuesarcomasarefibrosarcomasandinjectionsitesarcomas,
althoughothertypesofsarcomasdooccur.Fibrosarcomastendtodevelopontheheadandneck.In
youngcats,theyareassociatedwithFeLVandFSV.Injectionsitesarcomasdevelopincatsprimarilyat
sitesofpreviousvaccination,butdevelopmentfollowingothertypesofinjections(steroids)hasbeen
reported.SomestudiesincriminatethealuminumadjuvantintheFeLVandrabiesvaccines,but
whetherornotvaccineswithoutadjuvantaresaferisstillunclear.Currentguidelinesforvaccination
aretousetherearlegsofcatssinceamputationofthelimbispossibleifasarcomadevelops.Current
recommendationsfortreatingvaccinereactionsasgivenbytheVaccineAssociatedFelineSarcoma
TaskForcefollowsthe1,2,3rule;treatif1)themassisincreasinginsizemorethan1monthafter
vaccination,2)themassisgreaterthan2cmindiameter,or3)themassisstillpresentmorethan3
monthsaftervaccination.

Itisimportanttobeabletodistinguishavaccinereactionfromaninjectionsitesarcoma.Vaccine
andotherinjectionsitereactionstypicallyhaveamixofinflammatorycellspredominatedby
neutrophilsandmacrophages.Vaccinereactions,inparticular,canhaveaprominent
lymphoplasmacyticinflammatorycomponentaswell.Reactivefibroblasts,endothelialcellsand
histiocytescanmimicneoplasticsarcomacells;becautiouswhenyouseesuspiciouscellsona
backgroundofinflammation.Acutevaccinereactionsmayalsocontainadjuvantmaterial.This
materialispinktomagenta,andcoarselyclumped.Itcanbefoundextracellularlyorwithin
macrophagesandlookssimilartoultrasoundgelorlubricantmaterial.

Diagnosissarcomascanhavesomedifficulties.Sarcomascanbeconfusedwithgranulationtissue
orgranulomatousinflammationbecautiouswithinflammationandlookforverystrongcriteria
ofmalignancyorbiopsy.Sarcomasandpoorlydifferentiatedcarcinomascanappearsimilara
tumorthispoorlydifferentiatedshouldbeoverlymalignantinappearanceandbehavior.Sometimes
withcytologyallyoucansayismalignantneoplasiaandyoumaynotbeabletotellwhatkind.
Inflammationcaninducefibroblastandendothelialcellproliferation,becautiousinterpreting
pleomorphicmesenchymalcellspresentininflammatorylesionsunlessmultiplestrongnuclear
criteriaofmalignancycanbeestablished.Benigntumorssuchashemangiomaorfibromacanbe
mistakenforawelldifferentiatedsarcomaorreactivecells.Somecarcinomasinduceamarked
scirrhousresponseifonlythisportionofthelesionisaspirateditcouldbeconfusedwitha
mesenchymalneoplasm.

Histiocytoma
Thisisaverycommontumorofyoungdogs(1214%ofallskintumors)andtheoriginisthe
Langerhancellsoftheepidermis(histiocyticcells).Theytendtoformonthefronthalfofyoungdogs,
especiallytheheadandears,andmanywillspontaneouslyregresswithin3months.Althoughitis
classicinyoungdogs,Ihaveseenthemindogsofallagesandsometimesdogswillformmultiple
tumors.Theclassicappearanceisasmall,hairless,domeshapedlesionwhichmaybeulcerated,butI
haveseenthemdescribedasplaquelike.Oncytology,thetumoriscomposedofindividualized
roundcellswithvariablywelldefinedcellborders(sharptoindistinct)withvariablyshapednuclei:
round,ovalorslightlyindented(likeabinofpotatoes).Nucleihavefinechromatin,sometimes
nucleoliarepresentbutareusuallyindistinct.Cellshavemoderateamountsofpalestaining
17

cytoplasm;theouteredgesofthecytoplasmoftenstainlighterthantheperinucleararea,andthe
edgesmayberuffledinappearance.NOTE:youwillonlyseetheruffled,paleredgeifyouareina
thinregionwherecellscanspreadout.Anothercharacteristicfeature,whichyoumaysee,isthe
presenceofsmalllymphocytesscatteredinthebackground;thisusuallyheraldstheonsetof
regression.Histiocytomacellsareusuallyrelativelyuniform(littleanisocytosisoranisokaryosis)and
pale,anditisuncommontofindbinucleatedormultinucleatedcells.Occasionally,itisdifficultto
distinguishahistocytomafromaplasmacelltumor.

Mastcelltumor
Thisisaverycommonskintumorofdogsandcats.Theclassicfeatureisfindingmanymastcells,in
sheetsorindividualized.MCTrepresents16to21%ofallcanineskintumors.Approximately5060%
formonthetrunkand25%onthelimbs.Boxers,Bostonterriers,Labradorretrievers,beaglesand
schnauzersareatincreasedrisk,althoughboxerstypicallydeveloploworintermediategradetumors.
Tumorgrade,asassessedbyhistopathology,ishighlycorrelatedwithbiologicbehavior.Thereare
threegrades:high(undifferentiated,gradeIII),intermediate(gradeII)orlow(differentiated,gradeI).
Ingeneral,mastcellsthatareuniforminsizewithuniformnuclei,arewellgranulatedandhavesmall
uniformgranulesaremorelikelytobelowgrade.Thepopulationisveryuniformwithnobi,
multinucleatedorgiantcellsnoted.Poorlydifferentiatedcellscanbehighlypleomorphicwith
multiplecriteriaofmalignancy(giantforms,biormultinucleatedcells,markedanisocytosisand
anisokaryosis,visiblenucleoli,etc.).Theyalsotendtohavefewergranules(andoccasionalnoorvery
fewgranules)andthegranulesarelargerandmorevariablysized.Althoughcytologiccharacteristics
maysuggestloworhighgrade,itsimportanttorememberthatoneofthecriteriausedingradingis
invasionofsurroundingtissueswhichcantbeassessedoncytologythushistopathologyisalways
required.Lessthan10%oflowgradetumorsmetastasizewhile5596%ofhighgradetumorswill
metastasize.Preputial,scrotal,subungual,oralandmucocutaneousjunctionsitescarryahigher
gradeandworseprognosis.Lowgradetumorsareslowgrowing,solitary,rubberyandmaybe
hairlessbutusuallydontulcerate.Undifferentiatedtumorsarefastgrowingandmoreproneto
ulceration.SubcutaneousMCTsappearsimilartolipomasandcanbemistakenforthem.After
palpationormanipulation,youmayseeDarierssign:Erythemaandwhealswhichformdueto
releaseofvasoactiveaminesfromthegranules.

Clinicalgradeisalsoassociatedwithprognosis.GradingisbyRomannumeralfromItoIVwith
designationofabsence(a)orpresence(b)ofsystemicsigns.StageIaisbest,stageIVbistheworst.
Stagingisasfollows:
StageI:Onetumorconfidedtothedermiswithoutregionallymphnodeinvolvement
StageII:Onetumorconfidedtothedermiswithregionallymphnodeinvolvement
StageIII:Multipledermalnodules;largeinfiltratingtumorswithorwithoutregionallymphnode
involvement
StageIV:Anytumorwithdistantmetastasis,includingbloodorbonemarrowinvolvement.

Mastcelltumorsarethesecondmostcommonskintumorofcats(20%ofallskintumors).Most
cutaneousMCTsincatsarebehaviorallybenign.Thereisagradingsystemthatisdifferentfrom
dogswhichdividesfelinetumorsintocompact(welldiffereniated)ordiffuse(anaplastic);mostare
18

compact.Visceral(splenicorintestinal)MCTismuchmorecommonincatsvs.dogsandismuch
morelikelytoresultinwidespreaddisseminationtoliver,viscerallymphnodes,bonemarrow,lung
andintestines.Withsplenicforms,theremaybelongtermsurvivalwithsplenectomy,eveninthe
faceofwidespreaddissemination.Thesameisnottrueofintestinalformsthiscarriesagrave
prognosis.CutaneousMCTsappearassolitary,raised,hairless,wellcircumscribeddermalnodulesor
theycanbeflatandplaquelikeandresembleeosinophilicplaques.Someulcerate.Theyaremost
commonlyseenonthehead(ofteninvolvingthepinnaatthebaseoftheear)andneckfollowedby
thetrunkandlimbs.Inhorses,cutaneousMCTsareconsideredbenign.

Oncytology,sheetsofroundcellsareobserved.Welldifferentiatedcellsarefilledwithfinepurple
(metachromatic)granulesthatoftenobscurethenucleus,nuclearstainingisoftenpoor(theyoften
appearpalebluethegranuleshogallofthestain..).Granulecolordependssomewhatonthestain
used;DiffQuiktendstoproducesomewhatbluergranules,especiallyifyouareheavyhandedwith
thebluestain.Poorlydifferentiatedcellscanbehighlypleomorphicwithmultiplecriteriaof
malignancyandlowernumbersoflargergranules.Eosinophilicinflammationoften,butnotalways,
ispresent;itiscommonindogsandhorsesanduncommonincats.Reactivemesenchymalcellsand
collagenfibers(collagenolysis)maybeseen.Fibroblastscanbequiteprominentinsomecanine
tumors.Incats,smallwelldifferentiatedlymphocytesmayseenandinsomecatswithmultiple
tumors,thecellsmayresemblepoorlygranulatedhistiocytes.

Quickstains(DiffQuik)maynotstainmastcellgranules.IfyoususpectMCTbutarenotseeing
granules,tryawetmountprepwithnewmethyleneblue,itwillsometimesshowgranulesthatfailto
stainwithquickstains.Occasionally,MCTandgranularlymphomacanbedifficulttodistinguishfrom
eachother;bothcanoccurininternallymphnodesandorgans,andMCTcellswithscantcytoplasm
andfewgranulescanbeconfusedwithgranularlymphoma.Ifyouareconcernedaboutthis,havethe
slidesevaluatedbyaveterinaryclinicalpathologistandconsiderdoingadditionaldiagnostictests
(biopsy,PCRforantigenreceptorrearrangement[PARR]todetectclonallymphoidpopulations,etc.).

Plasmacytoma
Thisisanuncommontumorindogsandrareincats.Indogs,ittendstoformonthelips,earsand
digits.Thisistypicallyabenigntumorindogs,buttheliteraturesuggestsitmaybeamore
aggressivetumorincats.Cellsmaybewelldifferentiatedandeasilyrecognizedasplasmacellsorcan
exhibitprominentatypia.Cellsareroundwithdeeplybasophiliccytoplasmthattendstostaindarker
atthemarginsandpalerintheperinuclearzone.Thepaleperinuclearclearzoneisvariably
prominent.Nucleiareeccentricwithcoarsechromatin,andareusuallyroundbutrarelymaybe
beanorsickleshaped.Binucleatedcellsareacommonandconsistentfeature.Lownumbersof
multinucleatedcellsarealsofairlycommonandtypicallycorrespondtoaspirationofthecenterofthe
tumorwherethemorepleomorphiccellsarefound.Pleomorphismincaninecutaneoustumorshas
notbeenshowntocorrelatewithmetastaticpotential.

Lymphoma
Cutaneouslymphomamaybeprimaryor,rarely,partofdisseminateddisease.Cutaneouslymphoma
maybesolitaryorgeneralizedandisclassifiedasepitheliotrophicornonepitheliotrophic.
Extracutaneousinvolvementcanoccur.Indogs,Tcelllymphomaismorecommon.Appearanceof
19

thelesioncanbehighlyvariableandcanappearasnodules,plaques,ulcersanderythemicor
exfoliativedermatitis.Diseaseisoftenchronicbutoverallprognosisispoor.Incats,epitheliotrophic
formsaretypicallyTcellandnonepitheliotrophicformsareBcell.Lesionsoftenappearascrusty
plaquesandmaybepruritic.

Primaryskintumorscanhaveamorebizarreappearancecomparedwithlymphomaarisingin
lymphtissuesandcanalsohaveawaxingandwaningclinicalcourse.Cellscanvaryfromrelatively
smalltolarge.Nucleimayberound,indentedorevencerebriform.Nucleolimaybeindistinctor
prominent.Considerlymphomaifyouhaveanoddlookingroundcelltumoroftheskinthatdoesnt
fitanyoftheothercommonroundcelltypes.Thepresenceofsignificantnumbersof
lymphoglandularbodies(cytoplasmicfragments)shouldincreaseyoursuspicionoflymphoma
wheneverobservedinnonlymphoidtissuesororgans,includingperipheralblood!

ADDITIONALTUMORS&LESIONS...

Melanoma
Melanomaisatumorofmelanocytesandindogsandisuncommonincats.Inthedog,thisisa
commontumoroftheskinandoralcavity.Malignancyisbasedonlocation,histologyandbreed;
malignanttumorscarryaguardedprognosis.Tumorsofhairedskinareusuallybenign(85%).
Locationsassociatedwithmalignantbehaviorincludemucocutaneousjunctions(90%malignant),oral
cavity(90%malignant)andnailbed(50%malignant).Findingahighmitoticindexonhistologyis
predictiveofmalignantbehavior.Dobermanpinchersandminiatureschnauzersaremorelikelyto
havebenigntumors(75%benign);miniaturepoodlesaremorelikelytohavemalignanttumors(85%
malignant).Inthecat,histologicassessmentdoesnotappeartohavethesamepredictivevalueasin
dogs.Ingeneral,oculartumorsbehaveworsethanoraltumors,andskintumorscanbeeitherbenign
ormalignant.Ingrayhorses,melanomasarecommon.Theyformprimarilyonthetailbaseand
perineum,usuallystartingataround4yearsofage.

Cytologicappearanceofmelonomascanbehighlyvariabletheyareknownasthegreat
pretenderorthegreatimitator.Cellscanbeepithelioid,spindloid,ordiscrete/round.Themost
commonformsseenareepithelioid(looksepithelial)ordiscrete/round(cellsareindividualizedand
round).Poorlyornonpigmentedformscaneasilybemistakenforothertumortypesthusthis
tumor.Pigmentationvariesbothfromtumortotumorandwithintumors.Welldifferentiatedcells
haveabundant,finemelaningranuleswhichstaingreenblacktobrownblacktogoldenbrownto
blue.Heavilypigmentedcellsmayhavetheirnuclearfeaturesobscuredcellslooklikesolid
aggregatesofpigment+/apalespotwherethenucleussits.Poorlydifferentiatedtumorsmayhave
onlyafewcellswithaveryfinedustingofmelanin.Eveninwelldifferentiated,wellpigmented
tumors,cellsfromthedeepermarginstendtobelesswellgranulated.Featuresofmalignancywill
vary.Inlocationswheretumorsaremorelikelytobemalignant,malignancyshouldbepresumed,
evenifcellsareuniform,welldifferentiatedandlackovertcriteriaofmalignancy.Inlocationswhere
tumorsaretypicallybenign,findingcriteriaofmalignancyoncytologyincreasesthelikelihoodof
malignancy.

20

Therecanbedifficultiesindiagnosingmelanomas.Melanophagescanbemistakenfor
melanocytes,especiallyinlymphnodes.Amelanophagehasanoval,wellbehavednucleuswith
coarselyclumpedchromatin,andmelanintendstobeaggregatedincoarseclumpscontainedwithin
phagolysosomes.Melanocytestendtohavefinegranulesevenlydispersedinthecytoplasm.Poorly
pigmentedmelanomascaneasilybemistakenforothertumortypes,althoughmostwill,uponclose
inspection,haveatleastafewcellswithaslightdustingofmelanin.

Mammarytumors
Caninemammarytumorscanbecomposedofavarietyofelementsincludingmesenchymaland
epithelialcells,myoepithelialcellsandmatrixandtheymayallbeinthesametumor!Tumortype
dependsalotonwhatispresent.Becausecytologyspecimensmaynotsamplealloftheelements
thatarepresent,accuratelypredictingexacttumortypeisdifficult.Itisalsodifficulttodetermine
benignvs.malignant.Usingcellularcriteriaofmalignancy,about50%ofcaninetumorswouldbe
classifiedasmalignant.Buttheproblemisthatabouthalfofthosewillnotexhibitamalignantclinical
courseofbehavior(invasivenessandmetastasis).Sohowdowepredictwhichwillbebadactors?
Currently,thebestpredictorofmalignantbehaviorisassessmentofthetumorforevidenceoftissue
invasion;usingthiscriterioninconjunctionwithtumortype,about25%oftumorsareclassifiedas
malignantandthiscorrelatesfairlywellwithclinicalcourseofbehavior.So,arethereanyruleswe
canuseforcytology?Ingeneral,themorecriteriaofmalignancythatareseen,themorelikelyitisbe
malignant,butrememberthatsomebenigntumorscanhavesomeprettyuglylookingcellsinthem.
Thereversecanalsobestated;themoreuniformthecellsare,themorelikelyitistobebenign,but
histopathologyshouldstillbedone.Soiscytologyevenusefulforcaninemammarytumors?Itis
usefulforconfirmingamammarytumorvs.someotherdiagnosisanditcanbeusedtoscreenfor
metastaticdisease,butIwouldadvisehistopathologyovercytologyifyouknowyouaredealing
withamammarytumor.

Felinemammarytumorsarealittleeasier.Intact,cyclingcatscangetlobularmammaryhypertrophy
orfibroepithelialhyperplasia(oftendiffuse);bothshouldresolvewithspaying.Lobularhyperplasia
shouldbecomposedofuniformglandularepithelialcells.Fibroepitheialhyperplasialookslike
uniformspindlecellswithlotsofmatrix.Actualtumorsaretypicallyepithelialandabout80%are
carcinomas.Oncytology,carcinomasshouldlookepithelialwithcriteriaofmalignancy.Ifthecells
lookuniformandbenign,histopathologyisstillwarrantedtoruleoutacarcinoma.

Mucocele(AKASialocele)
Theaspiratedfluidfromamucoceleisclearorbloodyandoftenthickandstringy.Akeycytologic
featureisthebackgroundappearancewhichispinktobasophilicandoftenregionallydistributedin
somewhatthickclumpsorpuddlesofbluematerial.Inflammatorycellsareoftenentrappedwithin
thepuddlesofmucus.NOTE:thisappearanceisoftenbestappreciatedonlowpower.The
predominantcelltypeconsistsofindividualized,large,highlyvacuolatedmacrophages.Sometimes
theycanhaveanalmostspindledorstelateappearancebecauseofthetenaciousbackgroundfluid.
Nondegenerateneutrophilsarealsocommon,especiallyifthelesionhasbeentraumatized.Evidence
ofhemorrhagemaybeseen.

21

Dermatophytes
TrichophytonandMicrosporumspeciesaretheonlyspeciesofsignificantveterinarymedical
importance.Theseorganismsaredistributedworldwideandalldomesticspeciesaresusceptible.
Usually,theorganismsonlygrowonkeratinizedstructures(skin,hair,claws).

Oncytology,thearthrosporeshaveaboxylook;theyareroundtocuboidalandelongated,24m
indiamete,withathin,clearcellwallanddeeplybasophilicinterior.Youmayseethemadheredto
keratinflakesorhair,oryoumayseetheminvadinghairs.Inflammationmayormaynotbe
prominent,whenpresentitisusuallysuppurativetochronicsuppurativeandphagocytizedspores
maybeseen.Fewthin,septatehyphaemayalsobeseen.

Superficiallesionstypicallyconsistofflakingandcrustingwithhairloss,butcatscanhaveclinically
silentinfectionsandbeasourceofinfectionforotheranimals.Inimmunocompetentdogsand
shorthaircats,thisdiseaseisusuallyselflimitingandoftenoccursinyounganimals.Longhaircatsare
moresusceptible.Althoughinfectionisoftenlimitedtokeratinizedstructures,dogscommonly
developfolliculitisandfurunculosis.Catscandevelopgranulomatousdermatitisconsistingofwell
demarcateddermalnodules.Suppurativetopyogranulomatousinflammationistypicalandspores
and/orhyphaemaybeobserved.

22

NCSUVHCCYTOLOGYSERVICEINFORMATION:

NCSUCVMVHC

1060WilliamMooreDrive

ClinicalPathologyLaboratory,RmC269
Raleigh,NC27607

JaimeTarigo,DVM,DACVP
JenniferNeel,DVM,DACVP
CarolGrindem,DVM,PhD,DACVP
Email:VHCcytology@ncsu.edu
Phone:(919)7935082;(919)5136550
http://cvm.ncsu.edu/vhc/csds/Cytology.html

CytologySampleSubmissionGuidelines

Samplesmaybesubmittedforcytology,bloodsmearreviewandbonemarrowanalysis.
SampleseligibleforPARRwillbesubmitteddirectlytotheClinicalImmunology
Laboratoryuponrequest.

SlideSubmission:

Pleasesubmit6slidesmaximumforeachsiteunstainedwhenpossible.StainingoneslidewithDiff
Quiktocheckcellularityisrecommendedifcellyieldappearslow.
Pleaselabeleachslidewiththepatientnameandsite.

DROPOFForSELFMAILIN:Samplesmaybemailedtotheaddressabove.Forlocalhospitals,adrop
boxislocatedoutsideoftheAnimalScanEntranceoftheWellnessCenter(addressabove).

PREPAIDFedExMAILINOPTIONS:FEDEXcanbecalledforapickupatyourhospitalusingsupplied
prepaidlabels/envelopes(18004633339).Multiplesamples/patientscanbesubmittedinone
envelope.FedExOvernightShipping
$9(NC)$12(outofstate)

ReportingSchedule:
Cytologyreportsarefaxedand/oremailedwithin1to2businessdays.

Supplies
Pleasecontact(919)7935082forslideholdersandprepaidlabelsforovernightFedExshipping
envelopes.

Pricing
Cytology

BloodsmearReview
BoneMarrow
STAT

$30.00
$14.00
$40.00(submitslidesonlywithCBCreport,noEDTA)
$13additional(STATsampleswillbereadandreportedwithinthesame
dayreceivedpleasecall9195136550ifaSTATisneeded).

23

NCSUFLOWCYTOMETRY/PARRASSAYGUIDETOSAMPLESUBMISSION

SAMPLESUBMISSION
Whattypeofsampletosubmit:Submitasamplethatrepresentsthediseaseprocess.Seebelow.
ThesamplesaresimilarforflowcytometryandPARR.Forflowcytometry,weneedtoknowthe
species,andweneedasamplethatcanbemadeintoasinglecellsuspension.Ifthereareenough
cells,wecanrunbothassaysfromthesamesample.Noformalinfixedsamplesforflow!Formalin
fixedsamplesOKforPARR(atanadditionalcostseebelow).

Presentingcomplaint

Bestsampletosubmit

Formofsample
Lymphadenopathy

Lymphnodeaspirate

Tubew/saline
Lymphocytosis

Blood

EDTA
Suspectbonemarrowdisorder

Bonemarrowaspirate

EDTA
Mass

Aspirate

Tubew/saline
Effusionw/cells

Fluid

EDTA
CSFw/cells

CSF

EDTA

Howtosendasample:Refrigeratesampleimmediatelyaftercollection.Sendsamplewithacold
pack(besuresamplewillnotfreeze)forovernightdelivery.DonotsendforSaturday/Sundayor
holidayreceipt.SendbyFEDEX,UPS,orDHL.DonotsendUSpostal,asthedeliverywillbetothe
mainmailofficeoncampusandcantakeadayortwotoreachus.
SamplescollectedonFridaysmayberefrigeratedovertheweekendandsentoutthefollowing
Mondaymorning.
NOTE:Forblood,weabsolutelyneedCBCresultsfromasclosetotimeofsamplesubmissionas
possible.
FormalinfixedsamplesforPARR:3or42025micronsectionsina1.5mleppendorftubeor
equivalent(NOTONASLIDE).FormalinfixationcandegradeDNA,therefore,PARRmaynotprovide
pertinentinformationinapproximately10%ofthesecases.

Typicalturnaroundtimeforflowcytometryresultsis2448hoursafterreceiptofthesample.
ThetypicalturnaroundtimeforthePARRassayis23daysafterreceiptofthesample.

Noteaboutlowcellularitysamples:Somesamples(e.g.CSF)maynothaveenoughcellstorunflow
orPARR.Insomecases,wecandeterminethispriortostartingtheassays.However,oftenwedo
notknowuntilwehavecompletedtheassayandbeguntoanalyzetheresults.Inthesecases,asit
costsussameamounttorunasasamplewithenoughcells,youwillbechargedforthenon
diagnosticsample.

Prices:
Flowcytometry$90/sample;additionalsamplesatthesametimefromthesamecase
$55/additionalsample

PARRassay$105/sample(LN/massFNA,BM,effusions,blood),additionalsamplesfromthesame
caseatthesametime,$55/additionalsample
24

Formalinfixedsamples$130(asadditionalprocessingisrequired)CalltheClinicalImmunologyLab
(9195136363)priortosendingasample.Ifnooneanswersthephone,leaveamessage.Thisway
wecanbegintotrackthesampleifitdoesnotarriveinatimelymanner.Additionally,thereare
timeswhenweareshortstaffedortheUniversityisclosedandwecannotreceivesamples.Ifthisis
thecase,therewillbeamessagetothateffectonthephone.

25

JaimeTarigo,DVM,DACVP
JenniferNeel,DVM,DACVP
CarolGrindem,DVM,PhD,DACVP
VeterinaryHealthComplex
1060WilliamMooreDrive,RmC269 Email:VHCcytology@ncsu.edu
Raleigh,NC27607
Phone:(919)7935082;(919)5136550
Website:http://cvm.ncsu.edu/vhc/csds/Cytology.html

NCSUVeterinaryCytology

CytologySubmissionForm

Contact&BillingInformation

Veterinarian________________________________

Clinic______________________________________

Address____________________________________
City_______________State____Zip____________
Phone_________________Fax_________________
Email_____________________________________

PatientInformation
OwnerName_______________________________
PatientName_______________________________
Species:FELINE CANINE OTHER___________
Age_________Breed__________________________
GenderMN FS M F

Prepaido/nFEDEXLABEL
ShippingMethodDROPOFForSELFMAILIN*
*MailorDROPOFFtoaddressabove(DROPBOXlocatedatWellnessCenter/AnimalScanEntrance)

AdditionalTests
Cytology BloodsmearPathologyReview BoneMarrow

CytologySpecimenTissueAspirates,FluidSmearsOnly(contactforguidelinesforfluidsmearpreparation).

Date____________ STATREQUIRED? CALL(919)5136550TONOTIFY

#Sites__________(6slidesmaximumpersite)
Source(s)(Iflymphnode,pleasespecifylocation)
__________________________________________________

__________________________________________________

__________________________________________________
History
Clinicalpresentationincludingsymptoms,duration.VentralDorsal
__________________________________________________________________________________________

__________________________________________________________________________________________

__________________________________________________________________________________________

Grossappearanceoflesion(s)Whenapplicablelistsize/growthrate/mobility/texture/color.(Pleaseinclude

ultrasoundreportforinternalaspirates)_________________________________________________________
__________________________________________________________________________________________
__________________________________________________________________________________________
__________________________________________________________________________________________

NCSU FLOW CYTOMETRY/ K9/FELINE PARR ASSAY SUBMISSION FORM


Patient Information

Clinic Information

Animal name_______________________________

Clinic name________________________________

Owner last name____________________________

Clinic Address______________________________

Species/Breed_____________________________

_________________________________________

Age______________________________________

_________________________________________

Sex: FI_____ FS_____ MI_____ MC_____

phone_________________ Fax________________

SAMPLE TYPE

DATE COLLECTED

Aspirate: site_______________

______________

Flow cytometry______ PARR ________

Blood (EDTA)_______________

______________

Flow cytometry______ PARR ________

Bone Marrow (EDTA)_________

______________

Flow cytometry______ PARR ________

CSF ______________________

______________

Flow cytometry______ PARR ________

Other________________________

______________

Flow cytometry______ PARR ________

We will send your results via (check one) ______Fax

TEST REQUESTED

_____email (address____________________________)

**HISTORY** (IMPORTANT!!!!)
1) Signs or symptoms leading to test request (check all that apply)
___Lymphadenopathy (include most recent cytology report if available)
___Splenomegaly

____Hepatomegaly

___Lymphocytosis; lymphocyte number__________________(include most recent CBC/cytology report if available)


___Other peripheral blood abnormality (include most recent CBC/cytology report if available)
___ Bone marrow abnormality (include most recent cytology report if available)
___Mass; location_______________________ (include most recent cytology report if available)
___ Effusion/fluid containing suspicious cells; pleural_____ peritoneal____ CSF____ other_____________
___Hypercalcemia (value_____________)

____Hyperglobulinemia (value__________________)

2) Other signs and/or additional history or concurrent conditions including any treatment.
__________________________________________________________________________________________
__________________________________________________________________________________________

3) Is this patient an ehrlichiosis suspect? ____No ____Suspect ____Confirmed


Send overnight for morning delivery to: College of Veterinary Medicine
(Please use FED EX, UPS or DHL)
ATTN: Linda English, B-324
4700 Hillsborough St.
Raleigh NC 27606
NOTIFY LAB BY PHONE OR EMAIL PRIOR TO SENDING SAMPLE

Questions?
Clinical Immunology
Phone: 919-513-6363
Fax: 919-513-6703
email: Linda_English@ncsu.edu