Virology prac Flashcards
Describe the principle and method for use of fluorescent antibody staining for the detection of viral infections
- Cells infected w/ Japanese Encephalitis Virus (JEV) & is fixed: permealises membrane
- Diluted (1:10) Pt serum/plasma & incubated w cells: Aby enter cell & attach on JEV Ag in cytoplasm
- Wash off non-specific proteins
- Incubate w/ 2dary FITC-conjugated anti-Human Aby
- Wash, mount & view under fluorescent microscope
* JEV Ag in cytoplasm fluoresce green
* Always do Pos & Neg controls
P1: Understand the requirements for the growth of cells in culture.
Incubated at 37C in CO2
Distinguish primary/diploid and continuous cell cultures
Diploid: Normal cells (human/animal tiss). Mix of differentiated cells. Limited passage (/#replications?). Broad growth spectrum for viruses. e.g. human fibroblast
Continous: Mlaignant (human/animal). Use of particular cell lines useful. Indefinite passage. e.g. Vero cells
P1: Understand the application of PCR for the detection of viruses. Eg HPV
Primer directed against L1 region of HPV = detect 450bp fragment of L1 region of HPV genome
a) What does it mean PCR HPV screening have a high neg predictive value
b) Interpretation of PCR results for HPV screening & what it means
a) neg result the accurately not have virus
b) Neg: another check up in 5yrs
Pos: further testing on cytology OR sequence to determine risk type OR do LPC? (high quality)
Interpret these results if the pt’s of these samples are infected
a) IgM & IgG neg
b) IgM neg, IgG pos
c) IgM & IgG pos
a) not infected by virus in past/current OR lag phase
b) Had a past infection (IgG)
c) Had a recent (IgM) & past (IgG) infection
in fluorescent Aby staining, what preparation is required for IgM determination
sample must be treated to remove interfering IgG Aby by:
- ultracentrifugation
- Chromatography
- immunoabsorption (use anti-IgG Aby)
