Diagnostics Flashcards
What is a CRP test?
C-reactive protein
What anticoagulant is found in a red top tube?
none
What anticoagulant is found in a yellow top tube?
Gel to speed up clotting
What anticoagulant is found in a purple top tube?
Potassium EDTA
Haematology
What anticoagulant is found in a grey top tube?
Fluoride oxalate
poison
What colour tube would be used to measure U&E?
Serum in a yellow/red topped tube
What colour tube would be used to measure glucose?
Plasma in grey topped tube
What does HBA1c measure?
It is a long term control which shows damage to cells
if you are diabetic and have not complied with treatment until the day before your hospital appointment
What colour tube would be used to measure TFT? (thyroid function)
Serum in a yellow/red topped tube
What colour tube would be used to measure LFT?
Yellow/red topped tube
What is the difference between serum and plasma?
Serum is the yellow fluid produced when blood clots and you centrifuge it. All the clotting factors have been used up
Plasma is prior to clotting and still has clotting factors
Why is a blue tube used to measure clotting factors?
Has citrate which is an anticoagulant, stops red cells using calcium
Used to test clotting factors by adding calcium and measuring clotting time
What is added to a grey topped tube? What does this do?
Fluoride oxalate
It prevents the red cells from using glucose which they would otherwise consume, skewing the results
What enzymes are measured in a liver function test?
1) alkaline phosphatase
2) aspartate amino-transferase (AST)
3) alanine amino-transferase (ALT)
4) gamma glutamyl transferase (GGT)
What hormones are commonly measured in a hormone assay?
Thyroxine
TSH
Cortisol
Low sodium and high potassium electrolyte results indicate what?
Adrenal failure (or haemolysis due to difficult venepuncture)
What happens to urea and creatinine results in renal failure?
The rise
If a patient has high urea and low creatinine what would cause this?
Dehydration
What causes increased liver enzymes? Why?
Enzymes leak into the blood in liver disease.
Renal damage would show what urea and creatinine results?
Both would be high
What cardiac enzymes are measured?
Troponins
Creatine kinase (CK)
Aspartate amino transferse (AST)
Lactate dehydrogenase (LDH)
When do the cardiac enzymes rise following a heart attack?
CK- 4 hours
AST- the next day
LDH- 2 days later
What are the diagnostic methods for detecting a virus?
- Cell culture
- Electron micropscopy
- Antibody detection
serology- ELISA/EIA - Antigen detection
immunoflorescence- IF
enzyme immunoassay- EIA - Genome detection
PCR - Quantification of antibody or antigens
- Serotyping
- Quanitifcation of genomes- “viral load”
- Genome sequencing
What is sensitivity?
The test’s ability to correctly identify positive samples
What is specificity?
The test’s ability to correctly identify negative samples
What is serology? What is it tested on?
The detection of specific antibody or antigen in blood, CSF or saliva
What type of virology testing it conducted on throat swabs, nasopharyngeal aspirate, bronchoalveolar lavage and ET secretions?
IF or PCR
What type of virology testing it conducted on stool samples?
ELISA or PCR
What type of virology testing it conducted on urine samples?
PCR
What type of virology testing it conducted on clotted blood?
Serology (antibody detection
What type of virology testing it conducted on CSF?
PCR
What type of virology testing it conducted on blood (EDTA)?
PCR
Viral load testing
What type of virology testing it conducted on saliva?
Serology and or PCR
What antibody type is a marker for recent or acute infection?
IgM
What antibody is a marker for infection of immunisation in the past without the presence of IgM?
IgG
Viral isolation in cell culture is still needed for what condition?
Herpes simplex virus
How do you diagnose a respiratory virus?
PCR
What kind of sample is required for a respiratory virus?
- Throat swab +/- nose swab
- Nasopharyngeal swab
- Nasopharyngeal aspirate (NPA)
- Bronchoalveolar lavage (BAL)
- Endotracheal tube (ET) secretions
What samples are needed to diagnose a CNS disease, and for which test? (meningitis/encephalitis)
- CSF for PCR
- Stools and throat swab for PCR
- Blood for serology
What test is performed on a stool or vomit sample?
PCR
What are the common bacteriology diagnostic techniques?
1) Culture
- Sterile sites
- Non-sterile sites
2) Serology
3) Molecular techniques
4) Antimicrobial susceptibility testing
How do blood cultures display a positive result?
The bottom of the sample pot reacts with byproducts from bacterial growth causing a visible change in the colour of the pot
What condition is the only exception the rule that bacterial samples should be taken before antibiotic administration?
Meningitis
What colour do Gram-positive bacteria stain?
Purple/blue
What colour do Gram-negative bacteria stain?
Pink/red
What type of Gram-positive cocci typically form clumps when viewed microscopically?
Staphylococcus
How do you differentiate between the main groups of Staphylococcus bacteria? What do the different results diagnose?
Coagulase test
Positive: Staphylococcus aureus
Negative: Skin commensals
What type of coagulase test result would the type of staphylococcus that produced severe infections give?
Positive
What type of infections are caused by Staphylococcus aureus?
Infects skin/soft tissue
Endocarditis
Osteomyelitis
What type of infections are caused skin commensal staphylococcus?
Low pathogenic potential
Can infect prosthetic material causing line, pacemaker infections and endocarditis
What type of Gram positive cocci typically form chains when viewed microscopically?
Streptococci
What does streptococci tend to be sensitive to?
Penicillin
What type of bacteria are diagnosed by testing haemolysis?
Streptococci
What type of bacteria produce incomplete haemolysis? What is this result called?
α-haemolysis
Caused by:
- Streptococcus pneumoniae
- Streptococcus viridans
What type of bacteria produce complete haemolysis? What is this result called?
Complete haemolysis
Caused by:
- Streptococcus pyogenese (aggressive- strep. throat)
- Neonatal/paediatric infections
What types of bacteria commonly cause food poisoning?
- Salmonella (including S. typhi)
- Shigella
- Campylobacter
What bacteria would be suspected if a patient presented with diarrhoea containing blood or they were young/elderly?
E. coli O157
What bacteria would be suspected if a patient presented with diarrhoea that contained mucus or was green?
C. difficile
What bacteria would be suspected if a patient had recently had a holiday abroad or had rice-coloured stools
Cholera
What bacteria are routinely tested for in stool samples?
Salmonella
Shigella
Campylobacter
How is Clostridium difficile tested for in stool samples?
Toxin detection
OR
PCR for toxin gene
How are parasites tested for in stool samples?
Concentration and special stains
How does a lower prevalence affect the positive predictive value?
The lower the prevalence the lower the positive predictive value
What is the minimum inhibitory concentration?
The minimum amount of antibiotic required to inhibit the growth of an organism
What is the MIC breakpoint?
Minimum inhibitory concentration of an antibiotic
The obtained result from sensitivity testing must be higher than the MIC breakpoint
When does bacteraemia onset occur in relation to a fever?
Bacteraemia occurs half an hour before a fever
What are the basic symptoms of subacute bacterial endocarditis? How would this be tested for?
Symptoms: - Rash - Fever - Weight loss (+/-) Tested using blood culture
What are the basic symptoms of syphilis? How would this be tested for?
Symptoms: - Rash - Lymph nodes - Fever Serum is tested for antibodies
What are the basic symptoms of toxoplasma? How would this be tested for?
Symptoms: - Rash - Lymph nodes - Fever Serum is tested for antibodies
What are the basic symptoms of tuberculosis? How would this be tested for?
Symptoms:
- Fever
- Weight loss
Tested by culture and interferon γ
What are the basic symptoms of Brucellosis? How would this be tested for?
Symptoms: - Rash (+/-) - Lymph nodes - Fever Serum is tested for antibodies and blood culture
What antibody changes occur during the primary response to an infection?
After around 7 days is the peak of the IgM spike, then IgM declines and IgG takes over, peaking just before 14 days
What antibody changes occur during a secondary response to an infection?
After exposure to the antigen there is a huge IgG spike peaking at 12-14 days (1000 times the initial response)
There is also a small IgM spike, smaller that the spike from the first response
What tissue do histopathologists test?
- Biopsies
- Resection specimens
- Frozen sections
- Post-mortem
What do cytopathologists test?
Smears
Fine needle aspirates
What do histopathologists look for when testing biopsies?
Is it normal?
Is it inflamed, if so, what is the cause?
Is it cancer, if so, what type?
What do histopathologists look for when testing resection specimens?
How far has the cancer spread?
It all the cancer removed?
What are frozen sections used for?
Rapid freezing and sectioning process used during operations to determine if a patient has cancer and if it has all been removed
What is more common, hospital post-mortems or coroner’s post-mortems?
Coroner’s post-mortems
How are sections obtained for analysis?
Specimen must be properly labelled
Fixed in formalin
Embedded in paraffin wax
Cut into thin sections
What stain is typically used on tissue samples in histopathology?
Haematoxylin and eosin stain
What colour would healthy and sinister tissue be stained with a haematoxylin and eosin stain?
Healthy: pink
Sinister: purple
The more purple it is the worse it is
What tests are performed on sections in histopathology?
Stain (e.g. gram, Ziehl-Neelsen stain)
Identify specific antigens using antibodies (immunohistochemistry)
Carry out molecular tests
How long does a frozen section (histopathology) result take to reach the clinician?
30 minutes
How long does a biopsy (histopathology) result take to reach the clinician?
2-3 days
How long does a resection specimen (histopathology) result take to reach the clinician?
5-7 days
What are the different types of manufactured antibodies?
- Antisera from immunised animals (polyclonal)
- Monoclonal antibodies
- “genetically engineered” antibodies
How are monoclonal antibodes generated?
1) Mouse is immunised with antigen
2) Spleen cells (containing B cells producing antibodies to the antigen) are removed
3) Spleen cells fused with myeloma cells (immortal B cells) to form hybridomas
4) Culture in HAT medium then select for positive cells
5) Harvest monoclonal antibodies
How do you produce antibodies using recombinant DNA technology?
1) Isolate the population of genes encoding antibody variable regions
2) Construct fusion protein of V region using a bacteriophage coat protein
3) Clone a random population of variable regions which gives rise to a mixture of bacteriophages- a phage display library
4) Select phage with desired V regions by specific binding to antigen
What are the therapeutic uses of manufactured antibodies? Give examples
1) Prophylactic protection against microbial infection
e. g. IVIG, synagis (anti-RSV)
2) Anti-cancer therapy
e. g. anti-HER2
3) Removal of T-cells from bone marrow grafts
e. g. Anti-CD3
4) Block cytokine activity
e. g. anti-TNFα
What types of antibodies are used in drugs with the suffix “-omab”? Give examples
Mouse monoclonal antibodies
e.g. Muronomab, anti-CD3, transplant immunosuppression
What types of antibodies are used in drugs with the suffix “-imab”? Give examples
Chimeric or partly humanised antibodies
e.g. Infliximab (Remicade) anti-TNFα, Rituximab anti-CD20
What types of antibodies are used in drugs with the suffix “-umab”? Give examples
Human antibodies
e.g. Palivizumab, anti-RSV (Synagis)
What are the diagnostic uses of manufactured antibodies?
1) Blood group serology
2) Immunoassays
- hormones
- antibodies
- antigens
3) Immunodiagnosis
- Infectious diseases
- Autoimmunity
- Allergy (IgE)
- Malignancy (myeloma)
What is ELISA? How is it conducted?
Enzyme Linked ImmunoSorbent Assay
1) Add anti-A antibody covalently linked to enzyme
2) Wash away unbound antibody
3) Enzyme makes coloured product from added colourless substrate
4) Measure absorbance of light by coloured product
How does rapid testing (antibody) work?
1) Analyte added to strip with antibodies conjugated to gold nanoparticles
2) Capillary flow moves analyte along the membrane to antibodies which form complex if positive
3) Antibody/analyte complex reach the first test line and bind
4) Remaining antibody reaches second test line and binds
(If negative then no antibody/analyte complex will be formed so only second test line will show a result)
What is a common example of rapid antibody testing?
Pregnancy test
What is serum sickness?
Immune complexes in the circulation which deposit in the kidneys or joints
What is the immunological concern with a patient presenting with loss of appetite and weight loss?
Effect of poor nutrition on bone marrow cells
What is the immunological concern with a patient presenting with vague aches and pains?
Immune complexes
What is the immunological concern with a patient presenting with swollen glands?
Immune activation
What is the immunological concern with a patient presenting with fever, rash and small red patches, some lumpy?
Acute phase, activation, immune complexes
What is the process by which immune complexes cause damage?
Immune complexes can be small or large.
1) Vasoactive mediators cause separation of endothelial cells
2) Large complexes attach to the basement membrane; small complexes pass through the basement membrane and deposit around epithelial cells
3) Platelet aggregation occurs
4) Inflammatory cell damage
What are the different results that could be found from serum electrophoresis?
1) Normal healthy control would show dense band for albumin, then low circulating volumes of all other cells
2) Patient with an active immune response would show normal think albumin band plus a dense wide band showing increased immune cells (non-specific)
3) Could show a monoclonal expension of specific cell type (e.g. B cells) which could indicate a malignancy so would need to be investigated for myeloma
What are the different lymphocyte subpopulations?
- CD3+ T-cells (pan T cell marker)
- CD4+ T-cells (T helper/cells)
- CD8+ T cells (cytotoxic T cells)
- CD19+ B cells
CD56+ Natural Killer (NK) cells
How is flow cytometry performed?
1) A mixture of cells is labelled with fluorescent antibody with a different colour on each antibody
2) Stream of fluid containing antibody-labelled cells passed one at a time through a laser beam
3) Fluorescence and scatter properties are measured by the flow cytometer
Gives absolute quantification of cell types
What is the antiretroviral pathway for a patient with HIV?
1) Patient tested for antibodies to HIV
2) Perform CD4 count and viral load
3) Low CD4 count; high viral load
4) Commence ART (1st line therapy)
5) Every three months monitor CD4 count and viral load
6) CD4 count < viral load >
7) ART (2nd line therapy)
Increased life expectancy
What conditions arise as CD4 T cell count decreases in HIV?
500cells/μL: Bacterial skin infections, Herpes simplex, zoster; oral, skin, fungal infections
400cells/μL: Kaposi’s sarcoma
300cells/μL: Hairy leukoplakia; tuberculosis
200cells/μL: Pneumocystis pneumonia; cryptococcis; toxoplasmosis
100cells/μL: CMV; lymphomas
