Lab Exam 1 Flashcards
hemacytometer use
manual WBC, RBC, platelet counts and body fluid cell counts
depth and depth factor of hemocytometer
0.1 mm
10
area of each large square (9 total)
1 mm^2
area of each small square (25 in one large)
0.04 mm^2
counting area for WBCs
4 large corner squares
4 mm^2
counting area for RBCs
5 small squares in the center square
0.2 mm^2
counting area for PLTs
large center square (1 mm^2)
or 5 small squares if count in large square is >40
boundary line rules
- double line —count cells touching innermost top and innermost left side lines
- triple line—count cells touching innermost lines, and cells touching middle top and middle left side lines
WBC count dilution
1:20 with acetic acid
1.9 mL diluent. + 0.1 mL sample
RBC count dilution
1:200 with 0.9% saline
first, make 1:10 with 0.1 mL sample + 0.9 mL saline
then, make 1:20 dilution with 0.1 mL previous + 1.9 mL saline
PLT count dilution
1:100 with ammonium oxalate
the higher the dilution…
the more statistical error inherent in the procedure
formula for calculating cell counts
(# cells)(depth factor)(dilution factor)/counting area = cells/μ
when diluting for WBCs or PLTs, let sample sit for —- mins before charging hemocytometer to…
10 min
lyse RBCs
let cells sit in hemocytometer for —– min to…
10
let them settle into same plane
sources of error in manual count
- clumped/clotted/rouleaux
- chipped or scratched coverslip
- improper filling
- not mixing sample
- tech error when counting
normal RBC, WBC levels for an adult
RBC female: 3.8-5.2 x 10^6 cells/μL
RBC male: 4.5-5.9 x 10^6 cells/μL
WBC: 4.0-11.0 x 10^3 cells/μL
principle of manual hgb test
lysis of RBCs and conversion of hgb to methemoglobin, which is read on the spec
reagent, dilution, and wavelength of manual hemoglobin
SLS (sodium laurel sulfate)
1:250
540 nm
3 functions of Hgb reagent
- lysis RBCs
- dilutes blood
- converts hemoglobin to methemoglobin
Beer’s law equation for finding Hgb concentration
(ABS unk/ABS std)(std conc) = unk conc
Hgb sample dilution procedure
5 mL SLS
remove 20 μL
add 20 μL sample
manual Hgb interferences
icterus
lipemia
high WBC ct
high plasma proteins
failure to lyse cells
failure to detect abnormal Hgb
hematocrit
proportion of blood that is RBCs
spun crit centrifuge must be standardized every ——
6 months
degree to RBC packing during spun crit depends on…
- speed
- time
- radius of centrifuge
- fullness of tube (3/4)
- trapped plasma
advantages of spun crit
- uses small amt of blood
- only takes 3 mins
- allows us to look at supernatant
disadvantages of spun crit
- trapped plasma
- somewhat subjective
- incomplete separation of RBCs and buffy coat
causes of increased hematocrit
pregnancy
hemoconcentration
polycythemia/polycythemia vera
hypoxia → EPO → erythropoiesis
causes of decreased hematocrit
anemias
excessive fluids
many more
rule of 3s
Hct = 3(Hbg) ± 3
sources of error for spun crits
- hemolysis
- not sealed well
- IV contamination
- measuring buffy coat
optimum spin time
time at which maximum packing occurs
MCV
definition
RR
terminology
calculation
average size of RBCs
80-100 fL
normocytic, microcytic, macrocytic
MCV = 10(Hct/RBC)
MCH
definition
RR
calculation
average weight of Hgb in RBCs
28-34 pg
MCH = 10(Hgb/RBC)
baby MCV
up to 105
MCHC
definition
RR
terminology
calculation
hemoglobin concentration in average RBC
32-36 g/dL
normochromic, hypochromic, spherocyte
MCHC = 100(Hgb/Hct)
causes of falsely elevated MCHC
- lipemia
- icterus
- hemolysis
- high WBC
- high protein
- cold agglutinins
causes of falsely decreased MCHC
- hyperglycemic osmotic effect
- IV contamination
RDW
definition
obtained from…
RR
unable to report when…
RBC distribution width
obtained from RBC histogram
12-15 (CV)
unable to report with dimorphic RBC population
anisocytosis
associated with which index?
variation in cell size
RDW
should always be used to check the indices
appearance of cells on smear
H+H mismatch
MCHC <32
check glucose/Na/K
or contaminated specimen
recollect or spun Hct and report
H+H mismatch
MCHC >36, <40
MCV low/normal
spherocytes
check smear
report
H+H mismatch
MCHC >36
MCV normal
lipemia, icterus, high WBC count
saline replace
H+H mismatch
MCHC >36
MCV high
cold agglutinin
warm, or saline replace and warm
smear should be made when?
as soon as possible after collection
refrigeration makes cells more fragile
criteria of a good smear
- thick area makes gradual transition to thin area
- blood does not extend to end of slide; goes 3/4 way
- smooth with no ridges, spikes, holes
- rainbow effect on feathered edge
factors which affect thickness of smear
- angle of spreader
- speed of spreading
- size of drop
common causes of poor smear
- drop too large
- spreader pushed with jerky motion
- spreading too slow
3 purposes of diff
- screning
- diagnosis
- monitoring
Wright’s stain is a type of… (2)
polychrome stain
Romanowski stain
ingredients of wright stain and purpose
- methanol: fixative; no longer infectious
- eosin: acid dye, dyes basic components pink
- methylene blue: basic dye, dyes acid components blue/purple
eosinophilic
basophilic
neutrophilic
take up eosin
take up methylene blue
take up both
examples of eosinophilic substances
- Hgb
- granules of eosinophils
examples of basophilic substances
DNA
RNA
platelet granules
WBC nuclei
basophilic granules
pH of Wright stain
6.8-7.2
Wright stain excessively blue
pH too ↑
Wright stain excessively pink
pH too ↓
look for on feathered edge first
- WBC distribution
- platelet clumps
- fibrin strands
smudge cell procedure
if ≥9 are seen, do an albumin slide (1 drop albumin + 10 drops blood)
describe segs
- 10-15 microns
- clumped chromatin
- 2-4 lobes
- pink/tan granules
describe bands
- 1 lobed nucleus, usually indented
- chromatin through entire lobe
describe eos
- large orange circular granules
- 2-3 lobes
describe basos
- dark purple water-soluble granules
- light, smeared nucleus
describe lymphs
- round at rest
- large dense oval nucleus
- small blue cytoplasm
- countable granules
describe monos
- up to 20 microns
- more, blue-gray cytoplasm
- lacy chromatin
- folded nucleus
- vacuolated
basophil function
hypersensitivity rxn
IgE receptors
release histamine
WBC count frequently used by oncologists during patient chemo
ANC, absolute neutrophil count
-cytosis
increased
granulocytes, lymphs, monos
-cytopenia
decreased
granulocytes, lymphs, monos
-philia
increased
granulocytes
-penia
decreased
neutrophils
WBC estimate performed when…
analyzer’s WBC count is questionable
ie platelet clumps
WBC estimate procedure
- scan on 10X; find monolayer
- count WBCs in 1/4 field; do this for several, find avg
- average # is WBC count (x 10^3)
- if analyzer does not match w/i 20%, do a manual WBC ct
when is corrected WBC count for nRBCs done?
more than 5 nRBCs present per 100 WBCs
corrected WBC count for nRBCs
WBC(100)/nRBC + 100 = corrected WBC
platelet description
- small
- nonnucleated
- mainly consist of granules
younger platelets
report if you see many
giant platelets
RBC RR
WBC RR
platelet RR
female 3.8-5.2 x 10^6/μL
male 4.5-5.9 x 10^6/μL
4.0-11.0 x 10^3/μL
150-400 x 10^3/μL
platelet estimate performed when…
- platelet count critical (<30) for first time
- schistocytes present
- analyzer gives platelet population or platelet clump flags
platelet clumps may be due to…
EDTA sensitivity
not mixing when drawn
platelet estimate procedure
- on 100X, look at several fields on the feathered edge; check for clumps/clots (determines need for estimate)
- count number of platelets in at least 5 fields, and determine average
- multiply average by 15 and 20 to get estimated range
- if analyzer’s result is not within range, perform manual platelet ct
name and fix
giant platelet
“platelet F” method on analyzer
detects platelet organelles
name and fix
agranular platelets
“platelet F” method on analyzer
detects platelet organelles
report presence
name and fix
platelet clumps
recurs on same patient each day
EDTA sensitivity
draw in Na citrate
extra step when platelet count is done from Na citrate
multiply by 1.1 to account for additive volume difference
name and fix
platelet satellitism
EDTA sensitivity
draw in Na citrate
name and fix
fibrin strands
recollect
name and description
left shift
immature neutros in PB
infection response
name and description
toxic granulation
darker granules
toxic reaction to bacterial infection
name and description
dohle bodies
light blue areas in cytoplasm
leftover RNA
toxic change to neutro from infection
name and description
vacuolization
response to infection
result of phagocytosis
name and description
hypersegmentation
B12/folate deficiency
report if ≥5 cells with ≥5 lobes
name and description
agranular neutrophils
clear or blue cytoplasm
report
name and description
virally reactive lymphs
cytoplasm expands, wraps around RBCs
edges darker blue
name and description
plasmtacytoid cell
B-cell
eccentric nucleus
dark blue cytoplasm
if many are present, pathology follows up
name and description
large granular lymph
countable granules
NK cells or reactive T-cells
name and description
pediatric lymphocytes
appear younger
not abnormal
name and description
Pelger-Huet anomaly
genetic condition
bilobed (pince-nez shaped) nucleus
no effect on function
name and description
May-Hegglin anomaly
dohle-body like inclusions in granulocytes
large platelets and thrombocytopenia
genetic condition
name and description
Chediak-Higashi anomaly
giant granules in lymphs and neutros
fusion of granules
genetic
name and description
Alder-Reilly anomaly
large purple/red granules, very full
in almost all neutros
lysosomal storage disease (Hunter’s or Hurler’s)
name and description
smudge cells
broken cells
usually lymphs, especially in CLL
name and description
pyknotic cells
degenerated nucleus
solid mass
might be an old sample
