Brainscape's Knowledge GenomeTM

Browse over 1 million classes created by top students, professors, publishers, and experts.


See full index

Hem 240 > Retic analysis and Validation > Flashcards

Retic analysis and Validation Flashcards

1
Q

What does a pronormoblast have

A

Highn: C ration
Round nucleus with nucleoli
high concentration of ribosomes
lots of RNA
able to divide
-dark blue cytoplasm
-found in bone marrow
-biggest

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

What does a basophilc normoblast have

A

high N:C ratio
very dark blue cytoplasm
-able to divide
-found in the marrow
dectectable hgb synthesis
-lots of RNA and ribosomes
-dna synthesis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

Polychromatic Normoblast what does it have

A

-N:C ratio starts to even out
-pink cytoplasm that reflects complete hgb production
-last stage of cell able to undergo mitosis
-found in the bone marrow
-increased hgb synthesis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

Orthochromic normoblast

A

-low N:C ratio
-chromatin is condensed
-pink cytoplasm
-residual ribosomes present and RNA/Organelles are degrading
-start of cells not being able to divide
-loses nucleaus

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

Polychromatic erythrocyte

retic

A

-no nucleus
-no division
-moves from marrow to circulation
-residual ribosomes and organelles
-increased acidophilia
-no DNA because there is no nucleus

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

mature erythrocyte

A

-no nucleus
-no division
- found in circulation for 120 days until removed by spleen
-delivers O2 to tissues
-its biconcave shape helps for optimal O2 exchange
-flexibility allows it to enter small capillaries

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

Increased Retic count means what

A

hemolytic anemia
hemorrhage
treated IDA and megaloblastic Anemia
Uremia

-proper BM response

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

DECREASED RETICULOCYTE COUNT

A

Ineffective Erythropoiesis as seen in:
pernicious anemia
sideroblastic anaemia
Thalassemia
-Bone marrow failure so there are fewer precursors being released

Insufficient Erythropoiesis
Aplastic crisis
Aplastic anemia

decreased RBC production

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

Reticulocyte Staining

A

Wright’s Stain: Reflects Hgb present, residual ribosomes and RNA
Polychromatic erythrocyte

Supravital Stain:
Reticulocyte

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

MANUAL RETICULOCYTE COUNT
with supravital stain

A

-cells are alive
-New Methylene Blue or Brilliant Cresol Blue
-Reticulum is precipitated as a dye-protein complex
-The reticulum is RNA in ribosomes looks like a rope with knots you need more than 2 blue/black granules to call it a retic

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

Reporting retic count

A

report both relative in SI units and absolute in retics/L whole blood

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

SOURCES OF ERROR in retic count

A

refractive artefacts in RBCs caused by moisture in the air and poor drying

Poor mixing – retics float

Poor counting – missing or double counts

Blood and stain are not mixed before being made

Other RBC inclusions that stain supravitally can be mistaken for retics:
Howell-Jolly Bodies
Heinz bodies (stain at periphery)
Pappenheimer Bodies

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

Reticulocyte & Basophilic Stippling

A

both have RNA filaments

Reticulocytes:
“Rope with multiple tied knots” formation
the granules are uneven

Basophilic Stippling:
Fine or coarse granule evenly distributed throughout the cytoplasm
-unstable RNA that precipitated evenly

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

Corrected Retic Count

A

-in low Hct, the % retic may be falsely ↑ because whole blood contains fewer RBCs.
-A correction is made using the average normal Hct of 0.45 L/L

Assumption: Average HCT is 0.45 L/L

-dependent on anemia

  • Patients with 0.35 L/L to have elevated corrected reticulocyte count of 2-3% - That will tell us they’re compensating for their mild anemia
  • Patients with <0.25 L/L the count should increase to 3-5% to compensate.
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

Reticulocyte Production Index (RPI)

A

-Normal ‘retics’ mature in PB within 1 day
-Retics pushed to PB early due to a hemolytic anemia are called “shift” retics.
-Shift retics are released prematurely from the marrow to compensate for anemia itll take 2-3 for them to lose their reticulum

When assessing the BM response we check Retic count daily because retic count can be falsely increased when there is polychromasia

maturation time is based on a persons HCT
An RPI > 3 indicates a good marrow response
An RPI < 2 indicates an inadequate response
-indicates how good someones bone marrow response is

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

AUTOMATED RETICULOCYTE COUNT

A

-whole blood analyzed on a cell counter
-retics measured by flow cytometery; blood mixed with dyes or NA stain and the analyzer measure optical scatter or fluorescence
-auramine O is added to blood
-the more RNA the more fluorescence
-measures forward scatter which corresponds to cell volume/size and RNA content
-retics have high forward scatter and high fluorescence (high RNA count)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

Automated Retic Parameters

A

Immature Reticulocyte Fraction or IRF is replacing RPI
-ratio of immature reticulocytes to total reticulocytes in a sample
-the anaylzer can differentiate between immature retics that were released early and older ones
-newer ones have more RNA - BM response
-can be seperated into low, middle and high fluorescent pops
-so IRF is the sum of the middle and high pop since they have higher RNA wereas low pops are most developed since they have less RNA
-early indication of erythropoiesis and detect early therapy

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

how can IRF Absolute Retic Count be Used to distinguish diff types of anemias

A
  • Hemolytic anemias have high retic count, increased IRF
    -Chronic Renal Disease have decreased retic count, decreased IRF
    -Response to Nutritional Anemias have normal retic count, increased IRF
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

Reticulocyte Hemoglobin Concentration

A

To assess response to IDA treatment

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

Reticulocyte Indices

A

mean reticulocyte volume and distribution width

-early detection and diagnosis of iron deficient erythropoiesis in children

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

Quality Assurance in the Hematology Lab

A

Daily validation of methods & equipment

Detailed SOPs with sources of error

Reflex / confirmation testing

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

Validation of Operator

A

Licensed MLT
Member in good standing at CMLTO
Trained at an accredited MLS program
Michener
Properly trained on analyzer and proficient in the methods used for testing
Continuing Education

23
Q

Validation of Analyzer

A

Run & assess QC regularly
Manufacturer’s QC 1 / shift
Patient QC samples every ? Samples
Westgard rules – check system
Calibrate periodically
Preventive maintenance
Alerts for reagents & electronics

24
Q

Validation of Specimen:

A

Volume – Within 10% of stated draw volume
-Underfilled Tubes – FALSELY low RBC counts, HCT, morphological changes
-Overfilled Tubes – Impropoer mixing of anticoagulants . Resulting in PLT clumping or clots

Anticoagulant – Usually EDTA
-Acts as Ca2+ chelator (inhibits clotting), Preserves cellular components and morphology for assessment

Age/ Storage:
EDTA Blood Samples should be analyzed within 6hours of collection and kept at RT
-RBCs, PLTs swell as sample ages. WBCs degenerate

Mixing too aggressively may lead to hemolysis of RBCs&raquo_space; Messed up RBC parameters

25
Q

Validation of Results:

A

Rule of 3 – Normal HCT should be 3X of the Hgb +/- 3
-Failure to meet this may indicate sample integrity issues or abnormal RBCs

Linearity Limits – Results proportional to the calculated concentration/activity of the analyte
-Samples above linear range – Sample must be diluted and results need to be multiplied by dilution factor

Analyzer flags – Due to analyzer, specimen or patient abnormalities. Results must be investigated before release

Nonsense Results – Results are so out of range it’s seen as incompatible with life (sometimes due to interfering substances) MCV > 130, MCH/MCHC greatly increased

26
Q

Which parameters are directly measured?
Which parameters are calculated?

A

Which parameters are directly measured?
WBC Total Count and DIFF, RBC, PLT, Hgb
PLT volume/ MCV on new analyzers

Which parameters are calculated?
MCV on Michener AcT5diff AL analyzers
HCT L/L=[RBC(1012/L)x MCV (10-15L)]
MCH (pg)= [Hb (g/L)/RBC (x1012/L]
MCHC (g/L)= [Hb (g/L)/Hct L/L]
RDW-CV: derived from RBC histogram
WBC Absolute counts: derived from DIFF & Total WBC counts

27
Q

Impedance principle:
how the act5 works

A

How the analyzer counts cells

) Cells enter a conductive diluent
2) The counting chamber has a current applied between the external and internal electrode
3) When cell passes through aperture, this disturbs flow between internal and external electrode – Creates measurable voltage pulse
4) Oscilloscope displays pulses generated by the cells interrupting the current

Number of Pulses is PROPORTIONAL to number of cells counted

Size of Pulses is PROPORTIONAL to size of the cell

RDW – Histogram showing size distribution of cells counted

28
Q

AcV technology

A

Absorbance
AcT5 - Tungsten halogen lamp
New Technology- laser optical scatter
AcT5-
Forward scatter (FS): 0°to detect cell volume or size
Side scatter (SS): 90° to detect internal complexity (granules, vacuoles etc.)

Cytochemistry
Diff Fix contains Chlorazole black
Stains granules of Neutrophils, Lymphocytes, Monocytes & Eosinophils

Volume
Focused flow impedance- measures volume of cell (change in resistance as cell passes through flow cell aperture)
Dual focused flow - where cells line up

Has a 5 part differential
Cytochemical staining
Differential Lysis
Technologies of Absorbance
Technologies of Cytochemistry
Coulter Impedance Principle

29
Q

WBC Diffplot signaling

A

Based on Cell Size (Volume) and Complexity (Absorbance)

Lymphocytes
Small volume, low complexity/absorbance

Neutrophils
Medium volume, moderate complexity

Monocytes
Large volume, low complexity

Eosinophils
Medium volume, high complexity

X-axis= Absorbance
Y-axis= Volume

30
Q

Technologies
again in the act 5

A

Electronic Impedance – Counting WBCs, RBCs and PLTs

Cyanmethemoglobin Method– Hemoglobin Determination

AcV Technology – WBC Diff Determination

31
Q

Reagent in the act 5

AcT5diff Diluent

A

counting and differentiating blood cells
-composed of saline
-stabilizes cell membranes for counting and sizing

32
Q

AcT5diff Fix

A

Lyses erythrocytes.

Fixes leukocytes.

Differentially stains granules of monocytes, neutrophils, and eosinophils

33
Q

AcT5diff
WBC Lyse

A

Lyses red blood cells for the leukocyte count.

Used to differentiate basophils.

34
Q

AcT5diff
Hgb Lyse

A

Lyses red blood cells.

Used to determine hemoglobin concentration.

35
Q

AcT5diff
Rinse

A

Enzymatic solution with proteolytic action.

Used as a rinsing agent.

36
Q

First dilution/Hgb bath:

what could go wrong

A

A dilution is made, and a certain amount is removed and sent for dilution in RBC/PLT bath
Remaining dilu’n is used for Hemoglobin Determination
RBCs are diluted again and lysed with HB Lyse reagent
Hbg released and is converted to cyanmethemoglobin and measured at 540 nm

What could go wrong?
-Lipemia or High WBC Count – Both increase turbidity which affects spectrophotometric reading of Hgb

37
Q

RBC/PLT Bath

A

Amount removed from First dilution/Hgb bath is then:
Further diluted using Diluent reagent
RBCs and PLTs are counted and discriminated by electrical impedance (Coulter principle)

RBC count = All blips between 30-300 fL
PLT Count = All blips up to 18 fL

38
Q

RBC and PLT Count

A

Develop the RBC histogram, which is needed to obtain the HCT, MCV, and RDW results
MCH calculated from RBC count and HGB value
MCHC calculated from HGB value and HCT
Develop the PLT histogram, which is needed to obtain MPV results

39
Q

WBC/BASO Bath

False cell counts?

A

Total WBC count is performed
-lyses RBCs and specifically differentiates between basophils and other leukocytes by volume

Next basophils are counted
Resistance of the basophils to the acidic lytic reagent is the basis for the differentiation of the basophils from the other leukocytes

False cell counts?
-nRBCs – resistant to lyse and can be counted as WBCs

100-400 fL – WBCs
300-400 fL – Likely basophils

RBCs were lysed, PLTs are too small and won’t register on histogram (>20 fL)

40
Q

Falsely increased RBC count?

A

-Very high WBC – They’re included in counts from 100-300 fL
-Giant PLTs – Counted as over 30 fL

41
Q

Falsely increased MCV? MCV calculations req. HCT and RBC

Falsely decreased MCV?

A

Very high WBC – may affect HCT and RBC – interfering substances
-Very small RBCs (ex. Fragments >20 fL) – They aren’t counted as RBCs. So this can FALSELY INCREASE PLT counts
-RBC agglutination – Increases RBC size so that it’s excluded from RBC count

Falsely decreased MCV?
-Giant PLTs – counted as small RBCs

42
Q

Red Cell Distribution Width

A

Tight RDW, most cells are a uniform size. Decreased anisocytosis = 11.4%

Large RDW indicates a large difference between smallest and largest cells therefore increased anisocytosis = 27

43
Q

Mean Cell Volume (MCV)

A

MCV RI = 80-100 fL

Right Shift – Expect macrocytes
Left shift – Expect microcytes
Two Distinct Peaks – 2 populations of cells

44
Q

DIFF Bath

A

Whole blood aliquot is mixed in DIFF bath with AcT5diff Fix reagent and Diluent
-cells are lysed
-Differentially stains Lymphocytes, Monocytes, Neutrophils, Eosinophils
-Primary focused flow for impedance (volume)
Second focused flow for optical detection (absorbance + cytochem)

45
Q

Where does the 5-part WBC Differential come from

A

Combined results obtained from WBC/BASO bath + DIFF bath

46
Q

Interfering substances
Lipemia

A

Turbidity affects spectrophotometric reading
- Falsely Increased – Hgb, MCH
-the instrument will show abnormal histrogram
-Corrective Action – Plasma replacement

47
Q

Interfering substances
High WBC

A

Increased turbidity affects spectrophotometric reading
-Falsely Increased – Hgb, RBC count,
-Incorrect HCT
-Rule of 3 will not match
-Corrective Action – Manual HCT, Correct RBC count, Recalculate RBC indices, Dilute the sample

48
Q

Interfering substances
cold agglutinins

A

clumps exclude it from being counted as RBC
Falsely Decreases –RBCs, MCV
Falsely increases – MCHC
-has a grainy appearance
-instrument can show right shift on RBC histrogram
Corrective Action – Warm specimen to 37C + rerun

49
Q

Interfering substances
PLT Clumps

A

Falsely Decrease –PLT count
Falsely Increases – WBC count
-large clumps are counted as platelets

Corrective Action
IF PATHOLOGICAL - Redraw specimen in sodium citrate and multiply result by 1.1
Corrective Action IF COLLECTION ISSUE – Redraw sample in EDTA, the and do 8-10 inversions

50
Q

Interfering substances
Nucleated RBCs

A

Lyse resistant so it gets counted with WBCs
Falsely Increase – WBC counts because nucleated RBCs are large so they are counted at WBC

Corrective Action – Make PBF and count number of NRBCs per WBCs to correct WBC count

51
Q

Microspherocytes/fragments

A

These are counted with PLTs bc so small

Falsely Decreased – RBC count
Falsely Increased – PLT count

May see LEFT shift in RBC histogram

Corrective Action – Look at PBF and confirm PLT estimate, then possible a manual PLT count

52
Q

Rule of 3

A
  • Hct should be 3x the value of the Hgb (± 3)
    -3 x Hgb/1000 = Hct ± 0.03 L/L
    -for Normocytic/Normochromic erythrocytes
    -indicate abnormal RBCs or may be 1st indicator of error (sample integrity)
53
Q

Delta Checks

A

used more in clinical
-compares current with most recent
-values can stay the same unless there was major intervention
-if the delta check fails this can indicate analytical error or mislabeled sample
-must investigate , sample held until cause is found

54
Q

Test and what poik should be noticed
Reticulocyte count
Iron studies
Sickle screen
Hemoglobin Electrophoresis
Osmotic fragility
DAT
G6PD Assay

A

Reticulocyte count-Increased polychromasia

Iron studies - hypo/micro

Sickle screen -Sickle cells

Hbel - N/N anemia with targets, folded cells crystals +/- sickles

Osmotic fragility -Spherocytes

DAT-Spherocytes & polychromasia

G6PD Assay-Bite/Blister cells

Hem 240 (6 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2024 Bold Learning Solutions. Terms and Conditions