Lecture 25 - HLA Methods Flashcards
1
Q
What does flow cytometry measure
A
- size
- granularity or internal complexity
- relative fluorescence intensity
2
Q
What are advantages of flow cytometry
A
- more sensitive
- cell viability is less of an issue
- less subjective readings
- detects only IgG antibodies
3
Q
How does a 3 colour flow cytometry crossmatch work
A
- labels T cells
- labels B cells
- labels bound antibodies
4
Q
What are properties of secondary antibodies
A
- affinity purified
- F(ab) fragment
- Fc specific
- No crossreactivity with mouse or rabbit immunoglobulins
- must be titred for maximum sensitivity
- laboratory developed test
5
Q
What are interferences in FCXM
A
- IVIg
- thymoglobulin
- ritiximab
- campath
- auto-reactivity
- any other antibody that targets T or B cells
6
Q
What is luminex cytometry
A
- luminex cytometer measures 3 colours and allows determination of 500 individual parameters in one tube
- 500 polystyrene beads labeled with varying degrees of 2 colours
- third colour used to detect antibody or probe attachment to the bead
7
Q
What are advantages of luminex
A
- can measure many parameters in few tubes
- less expensive than standard flow cytometry
- methods available for antibody analysis as well as HLA typing
- cannot run cells (no XM)
8
Q
What is cPRA
A
- panel reactive antibody
- positivity of a patient sample against a panel of cells representitive of the donor population
- c stands for calculated since it cannot be done with bead testing
