INTRODUCTION TO THE HEMATOLOGY LABORATORY.pdf

HEMATOLOGY

1. HEMA311 WEEK 2 LECTURE
OVERVIEW OF THE HEMATOLOGY LABORATORY

2. HEMATOLOGY
comesfromtheGreekwords“Haima”and“Logos”
disciplinethatstudiesthedevelopmentanddiseasesofblood
Inthisfield,thefundamentalconceptsofbiologyandchemistryareappliedtothe
medicaldiagnosisandtreatmentofvariousdisordersordiseasesrelatedtoor
manifestedinthebloodandbonemarrow
Physiciansrelyonhematologylaboratorytestresultstoselectandmonitortherapyfor
thesedisorders;consequently,acompletebloodcount(CBC)isorderedonnearly
everyonewhovisitsaphysicianorisadmittedtoahospital

3. FUNCTIONS OF THE HEMATOLOGY LABORATORY
Medicallaboratoryscientists,medicallaboratorytechnicians,laboratoryassistants,
andphlebotomistsemployedinthehematologylaboratoryplayamajorrolein
patientcare
Theassaysandexaminationsthatareperformedinthelaboratorycandothe
following:
Establishadiagnosisorruleoutadiagnosis
Confirmaphysician’sclinicalimpressionofapossiblehematologicaldisorder
Detectanunsuspecteddisorder
Monitortheeffectsoftherapy
Detectminimalresidualdiseasefollowingtherapy

4. HISTORY OF HEMATOLOGY
1657: Describe worms in the blood by Athanasius Kircher
1658: Discovery of erythrocytes by Swammerdam
1674: Human erythrocytes was described by Anton Van Leeuwenhoek
1842: GuilioBizzozerodescribed platelets as “petites plaques”
1846: PMN distinguished from other leukocytes by Wharton Jones
1879: First complete classification of leukocytes by Paul Ehrlich
1902: Development of Wright’s stain by James Homer Wright
1920: Hematology was considered separate science from clinical pathology

5. THE HEMATOLOGY LABORATORY TODAY
UseofautomatedanalyzerstodifferentiateandenumerateRBC’s;althoughWright’s
Romanowsky-typestain(polychromatic,amixtureofacidicandbasicdyes),and
refinementsthereof,remainsthefoundationofbloodcellidentification
RBC,WBC,andplateletappearanceisanalyzedthroughautomationorvisuallyusing
lightmicroscopyexaminationofcellsfixedtoaglassmicroscopeslideandstained
withWrightorWright-Giemsastain
Thescientifictermforcellappearanceismorphology,whichencompassescellcolor,
size,shape,cytoplasmicinclusions,andnuclearcondensation

6. BLOOD AND ITS COMPONENTS
aspecializedconnectivetissue;averagevolumeis5liters(adult)
COMPOSITIONOFBLOOD:
Liquidportion:SerumorPlasma(55%)
Cellularportion:Erythrocytes,Leukocytes,andThrombocytes(45%)

8. GENERAL CHARACTERISTICS OF BLOOD
Invivobloodisredandisinthefluidstate(invitro:coagulatesin5-10minutes)
Color:
Arterialblood:brightred(oxygenated)
Venousblood:darkpurplishred(deoxygenated;wasteproducts)
Thickandviscous(3.5-4.5xmoreviscoustanwater)
Averagespecificgravityof1.055
AveragepHof7.40(7.35–7.45)
Maintainedbyexcretionofcarbondioxide,acid-basebalance
Makesup75-85mLperkilogramofbodyweight
Bloodvolumedifferswithageandgender:
Adultmale:5-6liters
Adultfemale:4-5liters
Newborn:250-350ml

9. FUNCTIONS OF BLOOD:
Respiratory
Nutritional
Excretory
Bufferingaction
Maintenanceofbodytemperature
Transportofhormones
Defensemechanism
Coagulation

10. PLASMA
Liquid portion that transports and nourishes blood cells
Provides coagulation enzymes that protect the blood vessel
CHARACTERISTICS:
Color: pale-yellow
Pink: slight hemolysis
Red: gross hemolysis
Yellow-brown: icteric plasma (due to increased bilirubin)
Transparency: clear ; if milky/turbid: Lipemia(increased lipids inc.TAG)
CONTAINS FIBRINOGEN
Contains 90% of water and 10% solutes (proteins, lipids, hormones, etc.)

11. ERYTHROCYTES
Shape:biconcave(discoidshapecellordiscocyte)
Color:Salmonpink
Averagesize:6-8um(approximatesize:7.2um)
Hemoglobin
MaincomponentofRBC’s;givesitsredcolor
Function is to transport oxygen (oxyhemoglobin) and carbon dioxide
(deoxyhemoglobin)
Foundinthecentralpallor/pallorarea(occupies1/3oftheRBC:normochromic)
Ifcentralpalloris<1/3ofthecell–increasedhgbconcentration
(hyperchromicRBC)
Ifcentralpalloris>1/3ofthecell–decreasedhgbconcentration(hypochromic
RBC)

12. ERYTHROCYTES
Anemialoss of oxygen carrying capacity of the blood
Due to decreased RBC count
Decreased Hemoglobin concentration
Polycythemia
Increased circulating RBC’s (increased RBC count)

13. RBC PARAMETERS
RBC count
Hemoglobin and hematocrit (H/H)
RBC indices
RBC morphology

14. RBC COUNT
RBCcountisthenumberofredcellsin1uLor1literofblood
Usedfordiagnosisofleukemiaandpolycythemia
Types:
ManualRBCcounting–visualcountingofredcellsusingahemacytometer;
requires1:200dilution
RBCthomapipet
Hemacytometerorcountingchamber(reportedinuLorL)
RBCdilutingfluid–isotonicsolution(RBCsretaintheirshape,matches
osmolalityofblood);exampleis0.85%NSS;lysesWBC’s
Automatedcellcounters
Coultercounter–basedontheprincipleofelectricimpedance

16. Methodiscalledasthecyanmethemoglobin(HiCN)methodgoldstandardfor
hemoglobindetermination
Reliesonaweaksolutionofpotassiumcyanideandpotassiumferricyanide,called
Drabkinreagent
Automatedanalyzers-ionicsurfactant(detergent)sodiumlaurylsulfatetoreduce
environmentalcyanide
HEMOGLOBIN

17. HEMATOCRIT
Hematocritisoftenreferredtoasthepackedcellvolume(PCV),volumeof
erythrocytesorreadingofpackedcells(pRBC)
RatioofthevolumeofpackedRBC’stothevolumeofwholeblood
Reportedas%orL/L
TWOMETHODS:
Macrohematocritmethod
Microhematocritmethod

19. RBC INDICES
Measures parameters of a single RBC; useful in the assessment of anemia
Mean cell/corpuscular volume (MCV)
Measures the volume/size of a single RBC
Reported in femtoliters(fL)
Mean cell/corpuscular hemoglobin (MCH)
measures the weight/mass of hemoglobin in a single RBC
Reported in pictograms (pg)
Mean cell/corpuscular hemoglobin concentration (MCHC)
measures Hgbconcentration in a single RBC; staining capacity of RBC’s and amount
of central pallor (g/dL)
Red cell distribution width (RDW)
based on the standard deviation of RBC volume and is routinely reported by
automated blood cell analyzers
Measures the degree of anisocytosis

21. RBC MORPHOLOGY
Uses a light microscope and a Wright-stained blood film
Parameters assessed are:
RBC diameter
Color or hemoglobinization
RBC shape
Cytoplasmic inclusions
Anisocytosis -general term used in hematology to denote an increased variation in
cell size
Poikolocytosis-general term for mature erythrocytes that have a shape other than
the normal round, biconcave appearance (variation in cell shape)

24. RETICULOCYTES
InaWright-stainedbloodfilm,0.5%to2.5%ofRBCsexceedthe7-to8-mmaverage
diameterandstainslightlyblue-gray
Thesearepolychromatic(polychromatophilic)erythrocytes,newlyreleasedfromthe
bonemarrow
arecloselyobservedbecausetheyindicatetheabilityofthebonemarrowtoincrease
RBCproductioninanemiacausedbybloodlossorexcessiveRBCdestruction
Stainedbymethylenebluedyes
Nucleic acid stains or vital stains, are used to differentiate and count these young
RBCs
Vital (or “supravital”) stains are dyes absorbed by live cells
Young RBCs contain remnants of ribonucleic acid (RNA) in its cytoplasm

26. REFERENCE RANGE FOR RBC PARAMETERS (ADULT)

27. LEUKOCYTES
Looselyrelatedcategoryofcell
typesdedicatedtoprotecting
theirhostfrominfectionand
injury
aretransportedinthebloodfrom
bonemarroworlymphoidtissue
Nearlycolorlessinanunstained
cellsuspension
Leukocytosis –increased WBC
count
Leukopenia–decreased WBC
count
Leukemia–uncontrolled
proliferation of WBC’s

28. TYPES OF LEUKOCYTES
Neutrophil
Band neutrophil
Eosinophil
Basophil
Lymphocyte
Monocyte

29. NEUTROPHILS
Aphagocyticcellswhosemajorpurposeis
toengulfanddestroymicroorganismsand
foreignmaterial
NEUTs,segmentedneutrophils,SEGs,
polymorphonuclearneutrophils,PMNs
Segmentedreferstotheirmultilobed
nuclei
Neutrophilia –increased neutrophils
Neutropenia –decreased neutrophils

30. BAND NEUTROPHIL
Bandsarelessdifferentiatedorlessmature
neutrophils
Pinkorlavender-staininggranulesfilled
withbactericidalsecretions
LEFTSHIFT:anincreaseinbands;signals
bacterialinfection

31. EOSINOPHIL
arecellswithbrightorange-red,
regularcytoplasmicgranulesfilled
withproteinsinvolvedinimmune
systemregulation
Eosinophilia –increased eosinophils
Eosinopenia–theoretically not used

32. BASOPHIL
Basophilsarecellswithdarkpurple,
irregularcytoplasmicgranulesthat
obscurethenucleus
Basophilgranulescontainhistamines
andvariousotherproteins
Basophilia–increasedbasophils
Basopenia–theoreticalandnotused

33. LYMPHOCYTES
Lymphocytesrecognizeforeignantigensand
mounthumoral(antibodies)andcell-
mediatedantagonisticresponses
Mostlymphocytesarenearlyround,are
slightlylargerthanRBCs,andhaveround
featurelessnucleiandathinrimofnon-
granularcytoplasm
Lymphocytosis–increasedlymphocyteswith
variantorreactivelymphocytes;often
associatedwithviralinfections
Lymphopenia–decreasedlymphocytes
associatedwithdrugtherapyor
immunodeficiency

34. MONOCYTE
Animmaturemacrophagepassingthrough
thebloodfromitspointoforigin
Macrophagescompriseminorcomponentof
peripheralbloodWBC
Theirtasksaretoidentifyandphagocytose
(engulfandconsume)foreignparticles
Assistthelymphocytesinmountingan
immuneresponsethroughtheassemblyand
presentationofimmunogenicepitopes

35. MONOCYTE
Monocyteshaveaslightlylargerdiameter
thanotherWBCs,blue-graycytoplasmwith
fineazuregranules,andanucleusthatis
usuallyindentedorfolded
Monocytosis–increaseinmonocytes,
Foundincertaininfections,collagenvascular
diseases,orinacuteandchronicleukemias
Monocytopenia–seldomused

36. WBC PARAMETERS
WBC count
WBC differential count

37. WBC COUNT
Technique is the same as RBC counting
Refers to the number of WBC’s in 1 ulor liter of blood
Requires a dilution of 1:20; diluent used is a dilute acid solution (lyses RBC’s)
Leukocytosis ->11,500/ uL
Leukopenia -<4,500 / uL

38. WBC DIFFERENTIAL COUNT
A technique where 100 WBC’s are counted; 50 WBCs or 200 WBCs can also be
counted
a peripheral blood film is used; blood film is also checked for RBC, WBC, and
platelet abnormalities
Reported as the relative count: % of specific cell (Example: Neutrophils –65%)

39. THROMBOCYTES/PLATELETS
Truebloodcellsthatmaintainbloodvesselintegritybyinitiatingvesselwallrepairs;are
themajorcellsthatcontrolhemostasis,aseriesofcellularandplasma-based
mechanismsthatsealwounds,repairvesselwalls,andmaintainvascularpatency
Are2-4umindiameter,roundtooval,anucleateandslightlygranular
Somehematologistsprefertocallplateletsas“cellfragments”–fragmentsfrom
megakaryocytes
Thrombocytosis–elevatedplateletcount;signalinflammationortraumabutconvey
modestintrinsicsignificance
Essentialthrombocythemia-raremalignantconditioncharacterizedbyextremelyhigh
plateletcountsanduncontrolledplateletproduction
Thrombocytopenia–lowplateletcount;commonconsequenceofdrugtreatmentand
maybelifethreatening

40. THROMBOCYTES/PLATELETS
Uncontrolledplateletandhemostaticactivationisresponsiblefordeepvein
thrombosis,pulmonaryemboli,acutemyocardialinfarctions(heartattacks),
cerebrovascularaccidents(strokes),peripheralarterydisease,andrepeated
spontaneousabortions
PLATELETCOUNT
Usespipetteandhemacytometer,althoughadifferentcountingarea,diluent,and
dilutionisusuallyused
plateletsarehardtodistinguishvisuallyinahemacytometer,andphasemicroscopy
providesforeasieridentification
Automatedbloodcellanalyzershavelargelyreplacedvisualplateletcountingand
providegreateraccuracy

42. REFERENCE RANGE FOR WBC AND PLATELET PARAMETERS (ADULT)

43. COMPLETE BLOOD COUNT (CBC)
isperformedonautomatedbloodcellanalyzersand
includestheRBC,WBC,andplatelets
SPECIMEN:
Appropriateanticoagulantandtube(EDTA),
freefromclotsandhemolysis,sufficient
volume,testedandpreparedforstoragewithin
theappropriatetimeframe
ACCESSION:maybeautomated,relyingonbar
codeorradiofrequencyidentification
technology,thusreducinginstancesof
identificationerror
FLAGS
Anindicationwhenoneoftheresultsfromthe
profilingisabnormal
A“reflex”bloodfilmexaminationisperformed

44. HEMOSTASIS AND COAGULATION
Mosthematologylaboratoriesincludeabloodcoagulationtestingdepartment
Primaryhemostasis-Itreferstotheroleofbloodvesselsandplateletsinresponseto
avascularinjury
Secondaryhemostasis/Coagulationpathway-Describestheactivationofaseriesof
coagulationproteinsintheplasma
Tertiaryhemostasis/Fibrinolysis-Finalstageofcoagulation;thirdsystemofenzymes
andcofactorsdigestsclotstorestorevesselpatency

45. TESTS FOR HEMOSTASIS
Platelet count
Mean platelet volume (MPV)
Prothrombin time (PT)
Activated partial thromboplastin time (APTT)
Thrombin time
Fibrinogen assays
D-dimer assays

46. ADVANCED HEMATOLOGY PROCEDURES
Bone marrow examinations –uses a bone marrow smear (from BM aspirate or
biopsy)
Cytochemicalstains
Flow cytometry –immunophenotyping
Cytogenetic analysis
Molecular diagnostics –PCR, DNA sequencing

47. SPECIAL HEMATOLOGY PROCEDURES
Glucose-6-phosphate dehydrogenase assay
Sickle cell solubility tests
Hemoglobin electrophoresis and HPLC
Erythrocyte sedimentation rate
Osmotic fragility test

48. MEDICAL TERMINOLOGIES

49. MEDICAL TERMINOLOGIES

50. END