Manual Methods—Bench Questions Flashcards
WBC dilution
1:20
3% acetic acid
1.9 mL + 100 uL
procedure for WBC estimate
average WBCs in 1/4 field on 10x
manual count if >20% discrepancy from analyzer
WBC correction for NRBCs
wbc(100)/(nrbc + 100) = corrected wbc
how closely should sides on hemacytometer match?
<25 cells: ±5 cells
>25 cells: within 20%
normal wbc ranges for adult and newborn
- adult: 4.3-10.0
- newborn: 5.0-30.0
causes of increased wbc
- infection
- leukemia
- pregnancy
- exercise
causes of decreased wbc
chemo
viral infection
myelodysplastic syndrome
plt dilution
1:100
ammonium oxalate
lyses rbcs, but not retics
area to count for plts
<200: 1 large square
>200: 5 small squares
normal plt
critical plt
- normal: 140-400
- critical low: <30
- critical high: >800
plt estimate procedure
average plts on 100x * 15 and 20 to give range value should fall in
why isn’t MPV calculated from manual count?
no way to calculate; derived from plt histogram
increased MPV
younger plts being released; thrombocytopenia
causes of thrombocytopenia
ITP
HIT
DIC
TTP
ALL
aplasic anemia
causes of thrombocytosis
IDA
CML
essential thrombocytosis
normal retic adult and newborn
- adult: 0.5-2.0%
- newborn: 2.5-6.5%
main causes of elevated retic
- hemolytic anemia
- blood loss
- thalassemia
- response to therapy for anemia
Miller Ocular calculation
cells counted out of 112/10 = %
corrected retic takes into account…
calculation
smaller pool of rbcs in pts with low hct
corrected retic = retics(hct/45)
retic index takes into account…
calculation
premature release of retics during anemia
retic index = [retics(hct/45)]/maturation time
shift/stress retic
retics released early when H&H are low to compensate
principle of sickle solubility
- saponin lyses rbcs
- Na hydrosulfite reduces released Hgb
- reduced Hgb S is insoluble in phosphate buffer, so suspension is cloudy
—–% sickling Hgb must be present for positive sickledex
10
sickling hemoglobins other than Hgb S
- Bart
- C Harlem
- C Georgetown
Hgb S mutation
Chromosome 11
glutamic acid replaced by valine
Hgb S disease & trait on electrophoresis
- disease: mostly Hgb S, some F and A2
- trait: 60% Hgb A, 40% Hgb S, small amount of F and A2
possible causes for false negative sickledex (2)
- pt too young, <10% Hgb S
- total Hgb <8 g/dL
sickledex procedure if Hgb <8
add 40 uL, not 20
seen on smear of sickle disease & trait
- disease: sickles, targets, nrbcs, HJ bodies, pappenheimers, poly
- trait: targets
best sample for OF test
Na heparin
why are OF samples incubated 24 hours?
detect subtle abnormalities in OF that would not show up immediately
normal beginning and ending OF hemolysis values
begins at 0.45%
ends at 0.35%
HS rbc defect
spectrin membrane defect
why do thal pts have ↓ OF?
targets and hypos can take in more water before lysing
procedure for making KB control slides
- negative control: normal male blood
- 1:20: 1 drop cord + 19 drops male blood
- 1:10: 1 drop cord + 9 drops male blood
6 drops saline + 4 drops control blood; make 20 thin slides
reasons for performing KB
- determine necessary RhIg dose
- check for thalassemia (moth-eaten HgbF distribution)
- check for hereditary persistence of Hgb F
methodology of monospot
latex particles or rbcs coated with Ag specific to heterophile Ab in dipstick
dipstick is immunochromatographic
samples for monospot
serum preferred
EDTA plasma acceptable if tested within 24 hours
is monospot specific for infectious mono?
yes
seen on smear of EBV pt
atypical lymphs, often >20%
sx of EBV and other infections that cause similar sx
fever, malaise, sore throat, lymphadenopathy
CMV
toxoplasma
can cause false negative monospot
children <12yo and 10-20% of adults
reflex for negative monospot
EBV serology
if negative, CMV serology
normal CSF rbc and wbc for adult and newborn
WBC
adult: 0-10
baby: 0-30
RBC
0 for both
tube order for CSF
- chemistry
- micro
- hematology
distinguish bloody tap from hemorrhage
- distribution of blood in tubes
- macrophages with hemosiderin/rbcs
- clot formation(tap)
- xanthochromic
causes of xanthochromia
- rbc degradation
- bilirubin
- increased protein
- melanoma pigment
lining cells seen in CSF
- ependymal
- choroid plexus
bacterial infection CSF effects on glucose, protein
↓ glucose
↑ protein
corrective action for nrbcs found in CSF
WBC correction formula
what do we do with CSF of ALL children?
look at cytospin smear regardless of WBC count, to check for blasts
cause for a BF to be rejected for count
clot
lining cells
- mesothelials (serous)
- synoviocytes (joint)
- ependymal/choroid plexus (csf)
- epithelials (amniotic)
synovial fluid cannot be diluted with…
acetic acid (clot)
components of semen analysis and normal values (8)
- pH: 7-8
- count: >20 mil/mL
- motility: >50% motile
- liquefaction: liquefied
- viscosity: not viscous
- volume: 1.5-5 mL
- morphology: <30% abnormal
- vitality: 3+ to 4+
no sperm
procedure
azospermia
fructose test; check for bilateral aplasia of seminal vesicles or obstruction of ejaculatory ducts
sites for BM collection
- posterior iliac crest (adult)
- tibia (child)
BM aspirate used for…
- cytogenetics
- flow
- molecular
- cultures
BM core biopsy used for…
- architecture
- cellularity
handling for each type of BM test
- aspirate smear (EDTA, bedside)
- flow (Na hep)
- genetics (Na hep)
- PCR (EDTA)
- culture (SPS, or Na cit for viral)
- biopsy (sent to histology in jar of fixative)
BM diff procedure
- count every other field
- count 200 cells (100/slide)
- differentiate nrbcs
- assess cellularity, megakaryocyte, presence of malignants, M:E ratio
normal BM diffs show…
pyramid form of full range of maturation
ME: 2:1 to 4:1
normal cellularity: 100-pts age, ± 10%
5-10 megakaryocytes per 10x field
2 conditions causing hypersegmented neutrophils
- megaloblastic anemia
- myelodysplastic syndrome
signs of wbc dysplasia
- hyper/hyposegmented
- agranular
- asynchrony
signs of rbc dysplasia
- multinucleated nrbcs
- fetal traits
- asynchrony
- karyorrhexis
- ringed sideroblasts
signs of plt dysplasia
- giant
- micromegakaryocytes
- agranular
leukemoid rxn
- left shift
- increased wbc
- toxic changes
CLL on smear
lymphs
smudgs
high wbc
myelodysplastic syndrome on smear/bm
- dysplasia
- PB cytopenia in 1+ lines
- BM hypercellularity
leukoerythroblastosis associated with…
myelophthisis (crowding out)
toxic changes to neutrophils
- dohle bodies
- toxic gran
- vacuolization
