HLA Laboratory Currently

Question 1

advantages of flow cytometry over CDC

Answer 1

• more sensitive than CDC
• cell viability is less of an issue; esp. for B cells
• less subjective readings
• detects only IgG antibodies
• MORE EXPENSIVE

Question 2

properties of secondary antibody

Answer 2

• anti-human IgG
• affinity purified
• F(ab)’2 fragment
• Fc specific
• no cross-reactivity with mouse, rabbit Igs
• must be titred for maximum sensitivity (optimized for low Ab conctns)
• lab developed test

Question 3

interferences in the FCXM

Answer 3

• IVIg
• thymoglobulin (rabbit polyclonal Ab to thymocytes)
• Rituximab (humanized anti-CD20)
• Campath (humanized anti-CD52)
• auto-reactivity in pt serum
• any other antibody that targets T or B cells

Question 4

Luminex principle

Answer 4

• measures 3 colours and allows determination of 500 individual parameters in one tube
• 500 polystyrene beads are labeled with varying degrees of 2 colours
• the third colour is used to detect antibody or probe attachment to the bead

Question 5

advantages of Luminex

Answer 5

• can measure many more parameters in fewer tubes (i.e. less processing/washing etc. of samples)
• this cytometer is a lot less expensive than standard flow cytometer
• methids available for antiody analysis as well as HLA typing

Question 6

disadvantage of Luminex

Answer 6

cannot run cells so can’t be used for crossmatch

Question 7

Describe antibody analysis with Luminex

Answer 7

• each polystyrene bead can be labeled with an individual HLA antigen
• the MFI (mean fluorescence intensity) of each bead is measured
• because the panel is large, almost all HLA antigens are included in one well

Question 8

T or F. The Luminex has no callibraton curve and is semi-quanitative

Answer 8

T

Question 9

describe the cPRA

Answer 9

• panel reactive antibody
• refers to the postivity of a patient sample against a panel of cells representative of the donor population
• ‘c’ = calculated
• when antibody testing moved to single antigen beads, it lost the ability to be representative of the population frequency of an antigen
• cPRA tools turn the antibody specificities into a cPRA value

Question 10

%PRA

Answer 10

based on the known frequency of antigens in the popln

Question 11

what does %cPRA tell clinicians?

Answer 11

sense of donor availability

cPRA = 5% good news! 95% = not so good news

Question 12

T or F. Luminex is slightly more sensitive than flow XM

Answer 12

T

Question 13

HSP registry

Answer 13

highly sensitized patient registry is for [atoent with (>95% cPRA)

• deceased donor HLA typing entered here

Question 14

KPD program

Answer 14

kidney paired exchange program
- managed by CBS
- renal recipient and donor pairs not compatible can register
- matched runs performed regularly to look for pairs and chains
- national program