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Applications of Flow Cytometry - Coggle Diagram
Applications of Flow Cytometry
Transplantation
HLA antibody detection
Responsible for hyper acute rejection
Complement fixing Ab (IgG)
Pre and post-transplantation
Flow methods
Polystyrene beads coated in HLA specific molecules
Known HLA typed cells
Flow cytometer or luminex system
Luminex is beads only
Flow beads + cells
Non-complement binding Ab also detected
Long-term graft survival
Abs present in historical sera but do not have them now
Poor graft survival
Body has ability to produce Ab
25% graft survival
IgG against HLA Class I or II
Poor prognosis
Differentiate between IgG and IgM
IgG more serious
IgM
Donor primed but no hyperacute rejection
More/stronger immunsuppressants
Techniques
Flow PRA (One lambda)
HLA Ag bound to beads
Known colour on bead so it can be gated
Beads separated on basis PE
Positive results determined by binding of anti-human IgG FITC
Only 8 beads tested at a time
Lots of tubes needed
Luminex system
Detect up to 100 beads at a time
More HLA Ag detected
Screening beads and ID tests available
Screening bead is not specific
Principle
3 x beads
1 All Infrared
1 All visible red
1 50/50
Each with HLA A1/2/3 on surface
Add beads to patient sera
Incubate at 22-25\(^o\)C in the dark under agitation
Dark - no bleaching of beads
Agitation - no clumping
Temp - optimum for opsonisation
Wash to remove unbound Ab
No background positivity
Add reporter - anti-human IgG-PE
2 more items...
HLA typing
Flow crossmatching of organs
CD34 analysis of haematopoietic stem cell transplant (HSCT)
Transfusion
Immunophenotyping
DNA analysis
Immunology