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Four-color flow cytometric analysis of peripheral blood donor cell chimerism

Metes, D and Logar, A and Rudert, WA and Zeevi, A and Woodward, J and Demetris, AJ and Abu-Elmagd, K and Eghtesad, B and Shapiro, R and Fung, JJ and Trucco, M and Starzl, TE and Murase, N (2003) Four-color flow cytometric analysis of peripheral blood donor cell chimerism. Human Immunology, 64 (8). 787 - 795. ISSN 0198-8859

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Abstract

Passenger leukocytes have been demonstrated to play significant roles in initiating and also regulating immune reactions after organ transplantation. Reliable techniques to detect donor leukocytes in recipients after organ transplantation are essential to analyze the role, function, and behavior of these leukocytes. In this report we describe a simple, reliable method to detect donor cells with low frequencies using peripheral blood samples. Detection of small numbers of major histocompatibility complex (MHC) mismatched cells was first studied using four-color flow cytometry in artificially created cell mixtures. By selecting the CD45 + population and simultaneous staining with several leukocyte lineage markers (CD3, CD4, CD8, CD56, and CD19), MHC-mismatched leukocytes were consistently detected in cell suspensions prepared from directly stained whole blood samples with a threshold sensitivity as low as 0.1%-0.2%. When the fresh peripheral blood mononuclear cells were separated by conventional Ficoll gradient purification, similar, but slightly lower levels of donor cells were detected. Blood samples obtained 1-5 months after liver, kidney, and intestine transplants revealed that the kind of organ allograft influenced levels and lineage pattern of the circulating donor cells. This procedure provided a simple and reliable method in determining early chimerism in transplant recipients. However, the detection of MHC-mismatched leukocytes of all lineages was much lower when frozen peripheral blood mononuclear cells were used. © American Society for Histocompatibility and Immunogenetics, 2003. Published by Elsevier Inc.


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Details

Item Type: Article
Status: Published
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Metes, Ddim4@pitt.eduDIM4
Logar, A
Rudert, WA
Zeevi, A
Woodward, Jjew7@pitt.eduJEW7
Demetris, AJ
Abu-Elmagd, K
Eghtesad, B
Shapiro, R
Fung, JJ
Trucco, M
Starzl, TEtes11@pitt.eduTES11
Murase, N
Centers: Other Centers, Institutes, or Units > Thomas E. Starzl Transplantation Institute
Date: 1 August 2003
Date Type: Publication
Journal or Publication Title: Human Immunology
Volume: 64
Number: 8
Page Range: 787 - 795
DOI or Unique Handle: 10.1016/s0198-8859(03)00113-7
Institution: University of Pittsburgh
Refereed: Yes
ISSN: 0198-8859
Other ID: uls-drl:31735062121011, Starzl CV No. 2195
Date Deposited: 08 Apr 2010 17:37
Last Modified: 13 Oct 2017 22:59
URI: http://d-scholarship.pitt.edu/id/eprint/5581

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