9 - Diagnosing Infections Flashcards
How to make a specific aetiological diagnosis of infection
1) Demonstrate organism, component or product
2) Isolate the micro-organism (“gold standard”)
3) Demonstrate a serological response
Solid phase assay
Immobilise antigen of interest on solid phase.
Wash with known antibody with fluorescent tag.
How could the lineage of tumour cells be determined?
With immunohistochemistry.
Stain with tagged antibodies against specific cell surface markers.
Capture assay
Immobilise antibody of known specificity on solid phase.
Wash with sample.
Wash with tagged antibody of known specificity.
Nucleic acid detection methods
1) Hybridisation
2) PCR
Hybridisation
Fluorescently-tagged DNA sequence added to ssDNA sequence.
If it hybridises, DNA sequences are complementary
In situ hybridisation
Fluorescently-tagged DNA sequence is added to tissue section.
EG: Detecting HER2 in breast cancer tissue
Using PCR to identify the causative agent of bacterial meningitis in children 1) 2) 3) 4)
1) Testing for Strep pneumoniae, Haemophilus influenzae, Neisseria meningitidis.
2) Reverse primers are the same, unique forward primers
3) Fragment lengths for each bacterium will be different.
4) Run fragments on a gel, can tell which bacterium is possibly present by length of fragment.
Issue with PCR identification of bacteria
All you can really say is that there is a DNA fragment of similar length to that of suspected orgamism
How to tell if a serological response is new or old
1)
2)
1) Test antibody titres at 2 points in time, and see if titres are rising. 4-fold titre rise is diagnostic
2) High IgM titres are indicative of recent infection
Anti-IgM assays
1) Known antigen fixed to solid phase. Wash with patient’s serum. Wash with anti-human tagged antibody
2) Anti-IgM antibody fixed to solid phase. Wash with patent’s serum. Wash with known antigen of interest. Wash with anti-antigen tagged antibody.
