Measurement of HLA Antibodies

Sam Coker Sam_Coker at pall.com
Fri Jun 15 11:21:44 EDT 2007


Dear All:

I just want to express my sincere appreciation to all those that responded to my
request  for information on flow cytometric methods for measuring HLA
antibodies. The response has been great and this is the best forum where people
are ready to help others, fantastic.   I have pulled together all the
suggestions which may also be useful to others in this field.  Robert
Sutherland, thanks as always for your help.

Best regards
______________________________________________________________________________________
I refer you to
UNIT 6.16      Flow Cytometric Assessment of HLA Alloantibodies (
I am not sure if the above address is my institutional address so if so
it wont work for people outside my institution. However you can probably
find CP cytometry in your library.

Current protocols in Cytometry.
http://www.mrw.interscience.wiley.com/emrw/9780471142959/cp/cpcy/article/cy0616/current/h

tml
Sam,

OneLambda offers a flow based alloantibody detection system consisting
of beads coated with HLA peptides for MHC class I and II, including
single antigen beads.  These kits are expensive, as they are largely
used for clinical transplantation cross-matching.  See their web site:
www.onelambda.com
_________________________________________________________________
Depending on your application, it is also possible to detect
donor-specific alloantibody using a panel of CD40L stimulated B cells of
known MHC specificities.  See our work:
http://www.blackwell-synergy.com/doi/abs/10.1111/j.1600-6143.2003.00637.
x

Hope this helps

Martin Zand

_____________________________
Martin S. Zand, MD, PhD
Associate Professor of Medicine, Microbiology and Immunology
Medical Director, Kidney and Pancreas Transplant Programs
Co-Director, Center for Biodefense Immune Modeling
University of Rochester Medical Center
601 Elmwood Ave - Box 675
Rochester, NY 14642
Vox:  585-275-4517
__________________________________________________________
 One Lambda (CA) has flow beads with known HLA antigens on their
surface. Work very well in flow setup.


John A. McIntyre, Ph.D, FRC Path, D(ABHI)
HLA-Vascular Biology Laboratory
St. Francis Hospital
1500 Albany Street, Suite 1109
Beech Grove, IN 46107
Tel: 317.782.7193
Fax: 317.782.6949
e-mail: john.mcintyre at ssfhs.org
_______________________________________
Hi Sam,
I work doing clincial research in the HLA lab at the Puget Sound Blood Center in
Seattle.
Are you looking for the existence of Class I and II antibodies or are you trying
to identify them?
We use One Lambda's flow beads for Class I and II PRA (panel reactive antibody.
This product contains many bead groups with either class I or Class II antigens
on the beads that are combined together.  This will tell you that you have
antibody and give you a % of beads that react with the antibody. The higher
percentage shows a broader reactivity of the antibody and/or the higher number
of antibodies are present.
One Lambda's single antigen beads will allow you to identify the antibodies
present to a larger extent.  Not all HLA alleles are represented but many are
there.
We also use lymphocytes from patients and donors to identify the presence of
antibody.  Generally speaking T cells express Class I and B cells express Class
I and II.

If you can give me some more information of what you are looking for, I can give
you some more detailed informations.

I'll be off work from noon today until Wednesday June 20th.

Sincerely,
Mary Gallagher, MT(ASCP), CHS
Puget Sound Blood Center
HLA/Immunogenetics Dept
921 Terry Avenue
Seattle, WA  98104
____________________________________________________________________
Hi Sam,
We used One Lambda's flow-based products.  http://www.onelambda.com/

We now use One Lambda's luminex-based products.



Barbara J. Masten, PhD, D(ABHI)
Associate Professor
University of New Mexico School of Medicine
Department of Pathology
915 Stanford Dr. NE, BMSB 333
Albuquerque, NM 87131
_________________________________________________________________________Hi Sam

we use BD´s FITC-labelled HLA class II DP, DQ DR and it works perfect.

Cheers
Mikael
______________________________________________________________________
No problem.  My lab uses these all the time for transplant patients.
Are you looking for overall levels of antibodies, or are you trying to
define specificities?  For overall levels, use the FlowPRA I and II.
For specificity analysis, the single-antigen beads work great!	Let me
know if you have any questions.

Paul R. Warner, Ph.D., D. (ABHI)
Associate Director
Immunogenetics/HLA Laboratory
Puget Sound Blood Center
921 Terry Ave.
Seattle, WA  98104
(206) 373-6609 Office
(206) 292-1860 Lab
(206) 484-7285 Cell
(206) 559-1289 Pager
_________________________________________________________________________________________
______________

http://www.onelambda.com/product/productline.asp?productlineid=3
_______________________________________________________________________
I used to work with the One Lambda PRA Screening beads - it has been a
while, but they were fun when I did them.
Suzie Dapprich
________________________________________________________________________

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