Sequence-specific analysis of microchimerism by real-time quantitative polymerase chain reaction in same-sex nonhuman primates after islet and bone marrow transplantation

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Abstract

BACKGROUND. Accurate and sensitive detection of microchimerism in nonhuman primates (NHPs) after hematopoietic cell transplantation is essential for monitoring cell engraftment, for evaluating the success of transplant protocols, and for expanding the utility of NHP in transplantation studies. Because limited sequences are available for NHP major histocompatibility complex polymorphic loci, methods that can accurately determine low levels of donor cells in recipients with same-sex bone marrow transplantation are essential. METHODS. Thirty-seven pairs of primers, 16 from monkey and 21 from human, were screened with cynomolgus DNA samples. Real-time quantitative polymerase chain reaction was developed for accurately determining low levels of donor-specific DNA in the peripheral blood of islet/bone marrow transplant recipients of same sex cynomolgus monkeys. RESULTS. A total of six sets of primer and Taqman® probe combinations were included in this study, which are the most informative primer and probe sets ever reported for cynomolgus monkeys. Three pairs of primers were chosen from exon 2 of the Macaca DRB1 gene and another three pairs were chosen from human HLA DRB1 and DRB3 loci. Three of the six primer-probe sets were also found to work well for baboon (Papio hamadryas) and rhesus monkeys (Macaca mulatta). Sensitivity of the assay ranged from 0.03% to 0.1%, depending on the primer-probe set and donor-recipient pair. The methods are reproducible with relatively low standard error and coefficient of variation. CONCLUSIONS. This method is an informative, practical and sensitive method for the determination of donor-specific cells in the peripheral blood of NHP recipients of bone marrow transplant.

Original languageEnglish
Pages (from-to)1677-1685
Number of pages9
JournalTransplantation
Volume84
Issue number12
DOIs
StatePublished - Dec 1 2007

Fingerprint

Chimerism
Bone Marrow Transplantation
Primates
Sequence Analysis
Real-Time Polymerase Chain Reaction
Tissue Donors
Macaca fascicularis
Macaca mulatta
HLA-DRB3 Chains
Papio hamadryas
Bone Marrow
Transplants
HLA-DRB1 Chains
Papio
DNA
Cell Transplantation
Macaca
Major Histocompatibility Complex
Haplorhini
Exons

Keywords

  • Microchimerism
  • Nonhuman primates
  • Real-time PCR
  • Transplantation

ASJC Scopus subject areas

  • Transplantation
  • Immunology

Cite this

@article{e1a9da1223464307b04d7fb3edf6daa4,
title = "Sequence-specific analysis of microchimerism by real-time quantitative polymerase chain reaction in same-sex nonhuman primates after islet and bone marrow transplantation",
abstract = "BACKGROUND. Accurate and sensitive detection of microchimerism in nonhuman primates (NHPs) after hematopoietic cell transplantation is essential for monitoring cell engraftment, for evaluating the success of transplant protocols, and for expanding the utility of NHP in transplantation studies. Because limited sequences are available for NHP major histocompatibility complex polymorphic loci, methods that can accurately determine low levels of donor cells in recipients with same-sex bone marrow transplantation are essential. METHODS. Thirty-seven pairs of primers, 16 from monkey and 21 from human, were screened with cynomolgus DNA samples. Real-time quantitative polymerase chain reaction was developed for accurately determining low levels of donor-specific DNA in the peripheral blood of islet/bone marrow transplant recipients of same sex cynomolgus monkeys. RESULTS. A total of six sets of primer and Taqman{\circledR} probe combinations were included in this study, which are the most informative primer and probe sets ever reported for cynomolgus monkeys. Three pairs of primers were chosen from exon 2 of the Macaca DRB1 gene and another three pairs were chosen from human HLA DRB1 and DRB3 loci. Three of the six primer-probe sets were also found to work well for baboon (Papio hamadryas) and rhesus monkeys (Macaca mulatta). Sensitivity of the assay ranged from 0.03{\%} to 0.1{\%}, depending on the primer-probe set and donor-recipient pair. The methods are reproducible with relatively low standard error and coefficient of variation. CONCLUSIONS. This method is an informative, practical and sensitive method for the determination of donor-specific cells in the peripheral blood of NHP recipients of bone marrow transplant.",
keywords = "Microchimerism, Nonhuman primates, Real-time PCR, Transplantation",
author = "Dongmei Han and Dora Berman-Weinberg and Kenyon, {Norma S}",
year = "2007",
month = "12",
day = "1",
doi = "10.1097/01.tp.0000290680.66025.b6",
language = "English",
volume = "84",
pages = "1677--1685",
journal = "Transplantation",
issn = "0041-1337",
publisher = "Lippincott Williams and Wilkins",
number = "12",

}

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T1 - Sequence-specific analysis of microchimerism by real-time quantitative polymerase chain reaction in same-sex nonhuman primates after islet and bone marrow transplantation

AU - Han, Dongmei

AU - Berman-Weinberg, Dora

AU - Kenyon, Norma S

PY - 2007/12/1

Y1 - 2007/12/1

N2 - BACKGROUND. Accurate and sensitive detection of microchimerism in nonhuman primates (NHPs) after hematopoietic cell transplantation is essential for monitoring cell engraftment, for evaluating the success of transplant protocols, and for expanding the utility of NHP in transplantation studies. Because limited sequences are available for NHP major histocompatibility complex polymorphic loci, methods that can accurately determine low levels of donor cells in recipients with same-sex bone marrow transplantation are essential. METHODS. Thirty-seven pairs of primers, 16 from monkey and 21 from human, were screened with cynomolgus DNA samples. Real-time quantitative polymerase chain reaction was developed for accurately determining low levels of donor-specific DNA in the peripheral blood of islet/bone marrow transplant recipients of same sex cynomolgus monkeys. RESULTS. A total of six sets of primer and Taqman® probe combinations were included in this study, which are the most informative primer and probe sets ever reported for cynomolgus monkeys. Three pairs of primers were chosen from exon 2 of the Macaca DRB1 gene and another three pairs were chosen from human HLA DRB1 and DRB3 loci. Three of the six primer-probe sets were also found to work well for baboon (Papio hamadryas) and rhesus monkeys (Macaca mulatta). Sensitivity of the assay ranged from 0.03% to 0.1%, depending on the primer-probe set and donor-recipient pair. The methods are reproducible with relatively low standard error and coefficient of variation. CONCLUSIONS. This method is an informative, practical and sensitive method for the determination of donor-specific cells in the peripheral blood of NHP recipients of bone marrow transplant.

AB - BACKGROUND. Accurate and sensitive detection of microchimerism in nonhuman primates (NHPs) after hematopoietic cell transplantation is essential for monitoring cell engraftment, for evaluating the success of transplant protocols, and for expanding the utility of NHP in transplantation studies. Because limited sequences are available for NHP major histocompatibility complex polymorphic loci, methods that can accurately determine low levels of donor cells in recipients with same-sex bone marrow transplantation are essential. METHODS. Thirty-seven pairs of primers, 16 from monkey and 21 from human, were screened with cynomolgus DNA samples. Real-time quantitative polymerase chain reaction was developed for accurately determining low levels of donor-specific DNA in the peripheral blood of islet/bone marrow transplant recipients of same sex cynomolgus monkeys. RESULTS. A total of six sets of primer and Taqman® probe combinations were included in this study, which are the most informative primer and probe sets ever reported for cynomolgus monkeys. Three pairs of primers were chosen from exon 2 of the Macaca DRB1 gene and another three pairs were chosen from human HLA DRB1 and DRB3 loci. Three of the six primer-probe sets were also found to work well for baboon (Papio hamadryas) and rhesus monkeys (Macaca mulatta). Sensitivity of the assay ranged from 0.03% to 0.1%, depending on the primer-probe set and donor-recipient pair. The methods are reproducible with relatively low standard error and coefficient of variation. CONCLUSIONS. This method is an informative, practical and sensitive method for the determination of donor-specific cells in the peripheral blood of NHP recipients of bone marrow transplant.

KW - Microchimerism

KW - Nonhuman primates

KW - Real-time PCR

KW - Transplantation

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JF - Transplantation

SN - 0041-1337

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