Histocompatibility Flashcards
T or F. HLa antigens are protein molecules
T
This class of HLA is expressed on all nucleated cells and platelets
Class I HLA
each different DNA sequence at a given locus codes for an _______
allele
define polymorphic
MANY PPL HAVE A DIFFERENT DNA SEQUENCE FOR THAT GENE
polymorphism for HLA molecules are focused around this region
peptide binding region
which chromosomes are HLA genes located on
6
T or F. One of each HLA gene is inherited maternally and pternally
T, A, B, C, and DR, DQ
T or F. HLA antigens are co-dominantly expressd
T
HLA is associated with several autoimmune diseases such as:
Celiac
Type 1 diabetes
MS
Ankylosing spondylitis
HLA is also associated with drug sensitivities
Abacovir
Carbamazepine
why does the HLA typing matter?
typing of the donor allows the potential for allocation according to degree of HLA match
HLA typing also helps to interpret antibody specificities and explain and interpret crossmatch results
traditional HLA typing
- serological CDC methodology using lymphocytes
- lymphocytes isolated and tested against numerous anti-sera
- approx. 300 sera are required for a full HLA typing!
the standard method for HLA testing now
CDC (complement dependent cytotoxicity)
- lymphocytes incubated with serum (1 uL volumes)
- addition of rabbit complement = cell lysis when Abs attached
- cell lysis/death observed
T or F. We test all MHC genes for transplantation
F! only classical HLA
complete HLA allele for DR15
DRB1*15:03
DR = antigen
B1 = gene
15 = antigen (DR15 serologically)
03 = third allele described
HLA inheritance
- chromosome 6
- one of each (A, B, C, DR, DQ) inherited maternally and paternally
- Mendelian inheritance
- HLA co-dom expressed
6 loci of interest fo HLA typing
A,B,C = I
DR, DQ, DP = II
these HLA loci are traditionally typed in solid organ transplantation
A, B , DR
these HLA loci are more routinely typed
HLA C, DQA, DQB, and DP
list the molecular techniques for HLA typing
- SSP
- reverse SSOP (including Luminex based)
- qPCR
- sequence based typing- SBT vs NGS
- Oxford Nanopore
SSP
- sequence specific primer
- primers specific for every allele of interest included
- series of primers used almost like antisera
- many primers required to cover common HLA alleles
this has replaced SSP typing methods
reverse SSO typing
- loci amplified and many probes used to detect the different alleles
- uses a Luminex instrument for read out
- can run in batches or single samples
rSSO workflow
- isolated DNA amplified
- DNA amplified with HLA locus specific primers
- PCR product biotinylated
- PCR product incubated with probe labelled beads
- Streptavidin-PE labelled conjugate is added
- PE fluorescent on the beads is measured
advantages of DNA-based typing vs serology CDC
- serology requires viable lymphocytes (dies in -50 weather)
- age of specimen and conditions of transport are much less important
- easier to create a primer for an HLA antigen than to screen antisera for HLA specificities
- DNA samples easy storage for repeat testing
- higher res/more accurate typing
- less subjective readings
pitfalls of DNA based typing
- lymphocytes still needed for crossmatching
- potential DNA contamination demands stringent lab set-up and training
- rare “null” alleles will be detected by DNA but not expressed on the cells
- new alleles are always being identified so software must be frequently updated
