Blood tests Flashcards
On what type of blood are tests performed on?
Venous blood
How is venous blood collected?
Venepuncture
What is a vacutainer?
Evacuated tube system used to perform venepunctures
Vacuum inside
Automatically extract the exact volume of blood needed
Who collects blood samples?
Phlembotomists
Junior doctors or nurses collect the blood if patient is feeling unwell
What is an advantage of using vacutainers?
Only one needle needed
Change from one vacutainer to the next
What is the procedure of collecting blood using vacutainers?
Appropriate vacutainer is pushed into the needle-holder
Blood flows into the tube as it fills the vacuum
Once filled, the tube is removed
Another vacutainer can be inserted into the same vacutainer
What system is used to organise the vacutainers?
Vacutainers are colour-coded according to which additives they contain
DIfferent additives react with the blood differently
What are the different additives added to the vacutainers?
EDTA - purple
Sodium or Lithium Heparin - green
Sodium citrate - blue
Sodium or Ammonium Oxalate - yellow
Sodium fluoride - yellow
What does the purple top represent?
EDTA
What does the green top represent?
Sodium or Lithium Heparin
What does the blue top represent?
Sodium Citrate
What does the yellow top represent?
Sodium or Ammonium Oxalate
Sodium Fluoride
What is EDTA used for?
Anticoagulant
Chelates calcium (enzyme cofactor essential in the clotting cascade)
Used in full blood count tests
What is Sodium or Lithium Heparin used for?
Anticoagulant
Inhibits thrombin formation
Used in chemistry tests requiring plasma samples
What is Sodium Citrate used for?
Anticoagulant
Precipitates calcium
Used in coagulation studies
What is Sodium or Ammonium Oxalate used for?
Anticoagulant
Precipitates calcium
Used for blood glucose measurement
What is Sodium Fluoride used for?
Inhibits glucose metabolism
Preserves glucose concentration
Used with Sodium/ Ammonium Oxalate for blood glucose testing
In which ways can blood testing differ?
Additives added
Part of the blood that is tested :
- whole blood
- plasma
- serum
What are the three main fractions of blood?
Whole blood
Plasma
Serum
What is the process of testing whole blood?
Contains both cellular elements and plasma
Blood is collected into a tube containing and anticoagulant like EDTA
To prevent clotting
What is the process of testing plasma?
Blood is collected into a tube containing an anticoagulant such as heparin or EDTA
The sample is centrifuged
The supernatant formed following centrifugation is plasma
Plasma = whole blood - cellular elements
What is the process of testing serum?
Blood is collected into a plain tube and allowed to clot
The sample is centrifuged
The supernatant following centrifugation is serum
As the blood was allowed to clot first, serum does not contain clotting factors
Serum = plasma - clotting factors
What is supernatant?
Upper liquid layer formed when a suspension is centrifuged
What is centrifugation?
Blood samples are spin in a centrifuge at G-force of 1000
When is a FBC requested?
A routine investigation
The most frequently requested laboratory test
Almost all patients will undergo these
Measure parameters to do with different components of the blood
What conditions do FBC monitor?
Infection
Anaemia
Diagnose and monitor haematological malignancies
Different cells in the blood are investigated to detect underlying conditions:
- Leukaemia
- Hodgkin’s disease
- Myeloma
How to undertake a FBC?
Sample: 4ml whole blood collected in an EDTA (lavender top) tube
Test: FBC is conducted on an automated haematology counter
What are examples of modern counters?
Fluorescence flow cytometry - count WBC
Electrical impedence - count RBC and platelets
Colorimetric assay - measure haemoglobin concentration
Describe the process of fluorescence flow cytometry
Blood sample is diluted in buffer
Labelled with a fluorescence marker that binds nucleic acids
As cells flow through a narrow aperture they are illuminated by a laser beam
Three signals are recorded for each cell:
- Forward-scatter: indicates cell volume
- Side-scatter: indicates cell content
- Side-fluorescence: indicates amount of DNA and RNA
Cells with similar physical and chemical properties form a cluster in a scattegram
Allows the differentiation of WBC
How could WBC differential scattergram be used for diagnosis?
Each dot represents one celll
Coloured ovals indicate cells with similar properties
Cell types can be distinguished and counted using these parameters
Immature WBC have high fluorescence due to high nuclear activity
High numbers of these cells could indicate leukaemia
Describe the process of electrical impedance
RBC are very poor electrical conductors
Can be used to enable their measurement
RBCs are diluted in an electrolyte solution
They are then passed in a stream through a narrow aperture across which electrical current is maintained
Individual cells create an increase in the electrical impedance of a size proportional to their volume
Describe the process of colorimetric assay
Sodium lauryl sulphate is used to lyse cells
Heme group of Hb is oxidised, and SLS hydrophilic groups bind to form stable coloured complex
Monochromatic light from an LED is absorbed by the SLS-HGB complexes
The absorbance is measured by a photo-sensor
Absorbance is proportional to the haemoglobin concentration of the sample
What is SLS (sodium lauryl sulphate)?
Common detergent used in cleaning products and laboratory applications
Lyses both RBC and WBC
Denatures proteins by disrupting non-covalent bonds and swamping proteins with negative charge
What parameters are used to measure anaemia?
Haemoglobin
Size of RBC
What are the types of anaemia?
Macrocytosis
Microcytosis
What is macrocytosis?
Raised red cell volume
Indicates
- Vitamin B12/ folate deficiency
- ethanol
- hypothyroidism
What is microcytosis?
Reduced cell volume
Indicates
- iron deficiency
- thalassaemia trait
What are ways to investigate WBC?
Can indicate infection and haematological malignancies
Number of WBC
Look at differential subset and determine which one is abnormal
Can look at the appearance of the WBC by blood films
What is microcytic anaemia?
Low RBC but normal WBC
Small RBC and low haemoglobin
To find out underlying cause - conduct more tests:
- iron deficiency test
- blood film - look at cells under the microscope
What is a blood film?
Look at blood under the microscope
Blood films are made by smearing a drop of anticoagulated venous blood onto a glass slide
How are blood films prepared?
Blood films are dried, fixed with methanol and stained
A common stain = May-Grunwald-Giemsa stain
Automated preparation in hospital haematology labs
Slides are checked to endure adequate staining
Systemic visual inspection using light microscopy
Who looks at blood films?
Laboratory scientists
Alert the on-call haematology doctors if serious abnormalities are identified
What is hypochromia?
Lack of the characteristic red colour of RBC
Only a rim of haemoglobin is present at cell peripheries
What is iron deficiency microcytic anaemia characterised by?
Microcytosis
Hypochromia
Should FBC results be conclusive?
The normal range is describe for where 95% of the healthy population lies
This means 5% of the normal population will lie outside the normal range
Therefore never look at blood test results in isolation - look in context with the clinical symptoms
What happens with abnormal results in FBC?
Abnormal results are usually assessed in more detail by making a blood film
A repeat test would be conducted if the initial abnormal result is suspected to be artifactual
What are liver function tests?
Collection of tests that measured the concentration of certain proteins and enzymes in the blood
Provide information about the state of the liver
How do we carry out liver function tests?
5 ml of venous blood or plasma
No undue delay in preparing it
Can be sampled at any time - no need for patient preparation
What are precautions that have to be taken when taking blood for LFTs?
Using a tourniquet during a venipuncture should be avoided if possible, as this can cause falsely raised albumin levels
Bilirubin is broken down during exposure to light, so the blood sample should also be protected from the light
Are raised bilirubin levels evident clinically?
No
Above 50 microm = jaundice evident clinically in sclera and skin
Why can bilirubin be raised?
Problem with liver function
Problem with excretion
May be raised in metastatic liver cancer
Why can albumin levels be low?
Low due to liver function disrupted by cancer
Liver synthesised albumin
Why can AST and ALT levels be raised?
Raised as the damage to the hepatocytes make the enzymes leak out of the cells into the blood
ALT can be highly increased with liver 5x normal
What are urea and electrolytes used to measure?
Used to measure kidney function
Levels are closely controlled by the kidneys
Any disturbance to the kidneys function will cause abnormal levels of these
Where are lab tests run?
In Clinical Biochemistry labs
Run on automated analysers
How do automated analysers work?
Formed by modules
Connected to carry out different types of assay
Use colorimetric assays
What is a colorimetric assay?
Utilise reactions involving a colour change which can be measured using spectrophotometry
Can be used to measure lipids, drugs, proteins and enzymes
Advantages of using automated analysers
High throughput testing - 600 tests/hr
Contactless ultrasonic mixing avoids sample contamination
How are the biochemistry tests accomplished?
Automated process
Blood samples are labelled and barcoded
The barcodes are scanned by the analysers and contain the information about which tests run
Some large laboratories contain tracks along which samples are moved around to various test stations
Why are urea and electrolytes tested?
Provide information about kidney function
How are urea and electrolyte testing carried out?
5 ml of venous blood - serum or plasma
Carry out colourimetric and ISE assays
How can urea levels be used to indicate pathology?
Raised due to
- impaired kidney function
- dehydration
- GI bleeding - digestion of own blood
How can creatinine levels be used to indicate pathology?
Impaired kidney function
Why does impaired kidney function raise the levels of waste products in the blood?
If kidney function decreases, excretion of waste products decreases and concentration in the blood increases
What is Na+ used for in the body?
Required for
Fluid balance
Membrane potential
Nerve and muscle function
What is K+ used for in the body?
Required for
Membrane potential
Nerve and muscle function
Small changes can lead to changes in heart rhythm
What is Cl- used for in the body?
Required for
Fluid balance
Acid-base balance
What can creatinine be used for?
Used to calculate an estimated GFR
Cockroft Gault Equation
For males multiply answer by 1.2
For females multiply answer by 1.04
For those of Black-African origin multiply answer by 1.18
