Principles and automation of biochemical testing Flashcards
The lab setting
Increased demand for testing and advancements in tech
General chemistry analysers
Immunochemistry analysers
Automation is key to throughput
Rapid throughput is required to deal with increasing demands
Automation supports high throughput analysis
Turnaround time
General chemistry systems
Commonly used techniques within these systems
Small bench top analysers - large freestadning analysers
Spectroscopy
A=absorptivity
c = concentration
l = path length
Absorbance of substance will vary based on wavelength
If (a) is known concentration can be determined by measuring absorbance
Cuvettes can be key
Normally standard path length
can be disposable or reusable
Different cuvettes have different ranges of wavelengths
general chemistry analyser tests
Sample often have a range of tests conducted
Automation aims to facilitate multiple assays on one instrument
Requires maintenance
Interfcae
Dedicated software for instruments
Require to define test parameters
defined by manufactures
Specimens
Blood spectrums - serum or plasma
Other may be also used
presentations to instruments
Tubes
Sample aliquoting and presentation
Primary sample - presented in machine
Separator gel - following centrifugation
Sample identification - barcoding
QC and calibration materials can also be presented to the machine
Priority samples - identified with ‘stat’ facility
Pipetting
Defined volume of sample must be transferred accurately from the sample tube to the reaction tube to undertake testing
Problems with fixed probes
Carry over - transfer of material from one sample to another
Haemolysed samples - integration of centrifugation and analysis steps make automated
Reactions
Reaction vessel often consist of cuvettes, system temp controlled
Sequence often follows standard format
Types of reaction analysis - end of point reaction, kinetic reactions
Reagents - can be prepackaged, replenished as part of equipment, carry over for reagents
Mixing - mixer probes facilitate mixing
Data
Types
Data printed out or transmitted to lab info systems
Immunoassays
Antibodies raised against the analyse of interest bind it within a reaction sequence that can be monitored
Types of labels - enzymes, chemiluminescent molecules, fluorescent molecules
Two main types of immunoassay reaction - competitive, sandwich
Competiivitive immunoassays
Use a limited concentration of antibody to produce a reaction in which the test antigen and a labelled antigen compete for a limited number of sites on a complementary antibody
sandwich assay
Unknown analyte binds to an antibody
Analyte is washed before a second labelled antibody is added and attaches to bound analyte
Detection of labelled antibody is directly proportional to the level of analyte
Enzyme linked ImmunoSorbent Assay (ELISA)
Simple automated immunochemistry systems may involve pipetting units for sample/ reagent pipetting, enabling automated ELISA
Most ELISA systems can be applied to most ELISA methods and are open platforms/systems - allow us of own reagents
Majority of immunochemistry analysers are closed systems
Immunoanalysers
Some aspects similar to general analysers
Larger volumes as lower concentration of analyte
Carry over can causes issues due to potentially2 wide ranges of concentration between samples
Integration and Tracking
Two strategies - link 2 or more analysers together, using a single sample presentation system, combine general chemistry and immunoassay units
Processing required for most samples before analysis
Sample preparation before analysis are pre analytical steps - centrifugation, cap removal, aliquoting, sorting
Analytical steps - process of analysis to produce results
CSF Xanthochromia assay to support detection of Subarachnoid Haemorrhage (SAH)
Can be used to support investigation into a suspected subarachnoid haemorrhage (SAH) following an initial negative CT scan
SAH involves bleeding into the space and often results from rupture of an aneurysm in arteries within the space
CT scans are usually a front line tool to investigate SAH
