Lecture 2 - Methods of Measurement

Question 1

Benefits of Automated Analysis

Answer 1

Speed
- allow more rapid processing of samples
- leads to cost efficiency
Precision
Greater number of analytes
Identification of potential abnormalities 'flags'
- technical 
- pathological
Safety
- closed sampling
- decreased exposure to infectious agents
Quality assurance
- barcode identification of sample
- decreases errors due to misidentification/misinterpretation

Question 2

Flags

Answer 2

Identify situations that may be of technical or pathological significance
Form part of the report on the run screen/printed report

Question 3

Examples of RBC Flags

Answer 3

Microcytosis
- cause: MCV lower than range and presence of small RBC
- possible action: verify RBC morphology on slide
RBC Agglutination
- cause: possible RBC or platelet clumps
- possible action: verify RBC morphology, recollect sample if present

Question 4

What methods is the counting of cellular elements in blood based on?

Answer 4

Electrical impedance

Optical detection

Question 5

Electrical Impedance

Answer 5

Cell counting and sizing are based on the detection and measurement of changes in electrical impedance (resistance) produced by a particle as it passes through a small aperture
Based on
- RBC being poor conductors of electricity
- certain diluents are good conductors
Blood is greatly diluted in a buffered diluent
Controlled flow rate of sample through specified aperture
Current applied between two electrodes surrounding aperture
Cells displace diluting fluid -> increase in electrical resistance

Question 6

Optical Detection

Answer 6

Laser light is used
Diluted blood specimen passes in a steady stream through which a beam of laser light is focused
As each cell passes through the sensing zone of the flow cell, it scatters the focused light
Scattered light is detected by a photodetector and converted to an electrical pulse
The number of pulses generated is directly proportional to the number of cells passing through the sensing zone in a specific period

Question 7

Principles of Flow Cytometry

Answer 7

The cellular light scatter patterns can be used to identify cells
Light scattered along the axis of the laser beam is known as forward scatter
Light scattered perpendicular to the axis is known as ‘side scatter’ or ‘orthogonal scatter’
Forward scatter is indicative of cell size
Side scatter is indicative of cytoplasmic complexity
- greater side scatter occurs when granules are present in the cells cytoplasm
- e.g. granulocytes have greater side scatter compared to lymphocytes

Question 8

Technical Considerations of Analysers

Answer 8

Accuracy
Precision
Linear range
Throughput
Reliability
Costs

Question 9

Policy, Purpose and Scope of Analyser Maintenance and Quality Control

Answer 9

Policy - to maintain instruments according to supplier’s protocol and to perform QC procedures
Purpose - to consistently produce accurate full blood counts with minimal delays and interruptions
Scope - the procedures are performed regularly and as required to prevent, and where necessary, correct instrument problems

Question 10

Quality Control

Answer 10

Control sample of known value assayed at the start and end of each shift
The results are recorded and represented in Levey-Jennings plots
Reviewed to assess the operation of the analyser

Question 11

Quality Assurance Programs/Proficiency Testing

Answer 11

Provide samples for analysis from multiple labs
Each lab analyses and informs provider of results
Collate results of all the participants and provide comparison