PMID- 18535170
OWN - NLM
STAT- MEDLINE
DCOM- 20080930
LR  - 20080827
IS  - 1535-3702 (Print)
IS  - 1535-3699 (Linking)
VI  - 233
IP  - 9
DP  - 2008 Sep
TI  - PCR-based methodology for molecular microchimerism detection and quantification.
PG  - 1161-70
LID - 10.3181/0802-RM-35 [doi]
AB  - Peripheral blood microchimerism after pregnancy or solid organ transplantation 
      has been widely studied, but a consensus on its detection has not yet been 
      adopted. The objective of this study was to establish a panel of reproducible 
      molecular polymerase chain reaction (PCR)-based methods for detection and 
      quantification of foreign cells in an individual. We analyzed length 
      polymorphisms generated by short tandem repeat (STR) and variable number tandem 
      repeat (VNTR) markers. Human leukocyte antigen (HLA)-A and -B polymorphisms were 
      detected by reference strand conformation analysis (RSCA). Class II polymorphisms 
      on HLA-DRB1 locus were analyzed both by classical PCR-sequence-specific primers 
      (SSP) and by quantitative PCR (Q-PCR). Also, sex-determining region-y gene (SRY) 
      gene allowed specific male donor discrimination and quantification by Q-PCR in 
      female recipients. Binomial statistical distribution analysis was used for each 
      molecular technique to determine the number of PCR replicates of each sample. 
      This analysis allowed the detection of the lowest detectable microchimerism 
      level, when present. We could detect microchimerism in more than 96% and more 
      than 86% of cases at levels as low as 1:10(5) and 1:10(6) donor per recipient 
      cells (DPRC), respectively, using Q-PCR for SRY or for nonshared HLA-DRB1 
      alleles. These techniques allowed as low as 1 genome-equivalent cell detection. 
      Lower levels (nanochimerism) could be detected but not quantified because of 
      technique limitations. However, classical PCR methods allowed detection down to 
      1:10(4) DPRC for HLA-DRB1 PCR-SSP. The clinical application of these techniques 
      in solid organ transplanted recipients showed microchimerism levels ranging from 
      1:10(4) to 1:10(6) DPRC after kidney or heart transplantation, and 1 log higher 
      (1:10(3) to 1:10(6) DPRC) after liver transplantation. In conclusion, the 
      standardization of molecular microchimerism detection techniques will allow for 
      comparable interpretation of results in microchimerism detection for diagnostic 
      or research studies.
FAU - Pujal, Josep-Maria
AU  - Pujal JM
AD  - Translational Research Laboratory, Institut Catala d'Oncologia, Hospital Duran i 
      Reynals, Avda Gran Via s/n, Km 2.7, 08907 L'Hospitalet de Llobregat, Barcelona, 
      Spain. jmpujal@idibell.org
FAU - Gallardo, David
AU  - Gallardo D
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20080605
PL  - Switzerland
TA  - Exp Biol Med (Maywood)
JT  - Experimental biology and medicine (Maywood, N.J.)
JID - 100973463
RN  - 0 (HLA-A Antigens)
RN  - 0 (HLA-B Antigens)
SB  - IM
CIN - Liver Transpl. 2008 Sep;14(9):1373-4. PMID: 18773547
MH  - Adaptation, Biological/genetics
MH  - Alleles
MH  - *Chimerism
MH  - Female
MH  - HLA-A Antigens/genetics
MH  - HLA-B Antigens/genetics
MH  - Humans
MH  - Male
MH  - Microsatellite Repeats
MH  - Organ Transplantation
MH  - Polymerase Chain Reaction/*methods
MH  - Polymorphism, Genetic/genetics
EDAT- 2008/06/07 09:00
MHDA- 2008/10/01 09:00
CRDT- 2008/06/07 09:00
PHST- 2008/06/07 09:00 [pubmed]
PHST- 2008/10/01 09:00 [medline]
PHST- 2008/06/07 09:00 [entrez]
AID - 0802-RM-35 [pii]
AID - 10.3181/0802-RM-35 [doi]
PST - ppublish
SO  - Exp Biol Med (Maywood). 2008 Sep;233(9):1161-70. doi: 10.3181/0802-RM-35. Epub 
      2008 Jun 5.