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Hematology > Units 10&11 - Instrumentation/Automation > Flashcards

Units 10&11 - Instrumentation/Automation Flashcards

1
Q

The Coulter Principle

A

Counting particles/cells by electrical impedence

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2
Q

Principle of Coulter electrical impedence counters

A
  • Particles in a conductive fluid with current running through
  • Particles are bad conductors
  • Particles are sent in single file, they interrupt the current and that increases resistance creating a voltage pulse
  • Amount of voltage pulses = cell count
  • Amplitude (height) of each pulse = cell size
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3
Q

Early Coulter counters

A

One type of cell counted at a time
(ex. RBC at 60-120 fL)
Dilutions done manually, either for RBC WBC or PLT

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4
Q

Later Coulter Multiparameter Instruments

A

Counted different cells in separate counting chambers
Machine made dilutions and directed cells to appropriate channels

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5
Q

Coulter Model S in 1960s

A

Later multi parameter Coulter instrument

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6
Q

Modern Analyzers using Coulter principles

A

Lasers - fwd & side scatter
High freq. probe for conductivity
Retic count added
Slide makers added
Stainers added

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7
Q

Types of specimen for heme analyzers

A

Whole blood - WBC, RBC, PLT
Body fluids

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8
Q

How do cell counts differ in body fluids from whole blood

A

Body fluids have LOW cell counts
Proportion of WBC and RBC often inverted

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9
Q

RBC cell counts performed in what kind of solution

A

Isotonic solution (saline diluent)
Dilution very high so only one RBC read at a time

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10
Q

What must be done if high amount of WBC due to leukemia when automated RBC count?

A

Mathematical correction

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11
Q

WBC cell counts performed how

A

Diluted in fluid that lyses RBC
Similar to hemacytometer method
Same channel used to measure hgb; WBC diluent contains cyanide reagent for cyanmethemoglobin method

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12
Q

PLT cell counts performed how

A

In the same channel as RBCs
Instrument separates populations as to size

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13
Q

1fL = xliter

A

10^-15 L

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14
Q

1fL = xmililiter

A

10^-12 mL

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15
Q

1fL = xmicroliter

A

10^-9 mcL

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16
Q

x axis and y axis of histogram for cell counts

A

X axis: size
Y axis: frequency/count

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17
Q

Usual units for cell volume

A

femtoliter

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18
Q

Uses of histograms

A

RBC:
MCV, RDW, Visible interferences like clumped PLTs

WBC:
3 part diff on low end models

PLT:
MPV

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19
Q

Coincidence

A

Multiple cells passing through aperture at same time

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20
Q

How do coincidences skew values

A

False increase pulse height
False decrease cell count

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21
Q

How to fix coincidences

A

Decrease aperture size
Hydrodynamic focusing
Instrument software that edits out bad pulses
Correction tables
Dilute sample

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22
Q

Hydrodynamic focusing

A

Forcing cells into single file

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23
Q

Coincidences more common in specimen with..

A

High cell counts

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24
Q

Thresholds

A

Setting upper and lower size limit of each cell type

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25
Q

How thresholds are set for RBCs

A

Separate RBCs and PLT

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26
Q

How thresholds are set for WBCs

A

Granulocytes - Largest
Monos
Lymphs - Smallest

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27
Q

Action levels

A

Ranges for which counts are acceptable

28
Q

Below action level

A

SLide review
Manual count

29
Q

Above action level

A

Dilute
Repeat
Multiply by DF

30
Q

Critical Values

A

Instrument result acceptable but has serious consequences for patient
Phone doctor immediately

31
Q

Instrument flags/Prompts

A

Abnormalities detected
MLS must investigate

32
Q

If two or more of the triplicate counts are not in agreement range –>

A

No result reported

33
Q

3 Part Differential

A

Histogram of:
Lymphs
Monos
Granulos

34
Q

Lymphocyte size after diluent applied

A

35-90fL

35
Q

Monos size after diluent applied

A

90-160 fL

36
Q

Granulos size after diluent applied

A

160-450 fL

37
Q

5 Part Differential

A

Scatter plot - Coulter VCS
Volume
Conductivity
Scatter
Diluent keeps them in near native state

38
Q

Which differential keeps the cell sizes in near native state

A

5 part diff

39
Q

Volume (VCS)

A

Direct current
Essentially impedence
Cell size in fL

40
Q

Conductivity (VCS)

A

High frequency current
Measures opacity based on N/C ratio, nuclear density, granularity

41
Q

Scatter (VCS)

A

Laser light scatter
Internal structure
Granularity
Surface Characteristics

42
Q

Components of multi parameter Coulter like instrument

A

Specimen loading bay and rocker
Dilutor, tubing, channels to route cells
Counting chambers for WBC RBC Hgb
Power supply

43
Q

Coulter Reticulocyte Method

A

In separate channels, RBCs mixed with new methylene blue and diluent
Retics counted and reported as absolute amount
Comparison of retic maturity on scattergram and PBS

44
Q

Coulter Reticulocyte Diluent

A

Contains acid to remove hgb, leaving mRNA to stain
Hypotonic to convert all RBC to spherocytes - removes error based on shape

45
Q

How are measured CBC values run

A

Triplicates

46
Q

What are measured CBC values

A

MCV
WBC
RBC
Hgb
PLT count

47
Q

What are calculated CBC values

A

MCHC
MCH
Hct
RDW
MPV
3 or 5 part diff
Percent/abs counts for each WBC type

48
Q

Which hct is higher? calculated or spun?

A

Spun because of trapped plasma between RBCs

49
Q

Errors with Coulter-like instruments

A

Carryover
False increase in hgb
increased MCV
decreased RBC

50
Q

How is hgb measured in CBC

A

Cyanmethemoglobin
Same as manual

51
Q

When is carryover a problem?

A

If next specimen has low counts

52
Q

Why would false increase in hgb occur

A

Elevated WBC
Elevated lipids
(false inc in absorbance)

53
Q

Why would increase in MCV and decrease RBC happen

A

Cold agglutinins
Rouleaux
(clumping)

54
Q

What would you do to a specimen that has an issue with its plasma?

A

Replace plasma with saline

55
Q

What occurs in specimen with too much EDTA

A

RBC shrinkage
Hct false decrees
MCV and MCHC decreased

56
Q

When would a specimen have too much EDTA?

A

Underselling a tube

57
Q

MCHC calculated formula

A

(Hgb) * 100 / Hct

58
Q

MCH calculated formula

A

Hgb*10 / RBC

59
Q

Hct Calculated formula

A

MCV * RBC / 10

60
Q

RDW formula

A

CV of RBC size distribution

61
Q

Sysmex hematology analyzers scatterplot axes

A

X -Side Scatter
Y -Side Fluorescence

62
Q

Techniques used by Sysmex

A

Absorption spectrometry - SLS hgb method
Hydrodynamic focusing = single file (no physical aperture)
Counts cells by direct current (impedance)
Fluorescent flow cytometry (stains DNA)
Radio frequency

63
Q

Absorption spectrometry - SLS hgb method

A

Separate channel on instrument for hgb
Diluent lysis RBC
SLS (sodium laurel sulfate) converts Fe++ to Fe+++ for methemoglobin
Methgb combines with SLS –> chromatin absorbs at 555 nm
Run is fast with NO CYANIDE

64
Q

Hydrodynamic focusing and impedence

A

Same as Coulter only no physical aperture

65
Q
A

Hematology (12 decks)

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