Amplification of a Y-chromosomal DNA sequence by the polymerase chain reaction for documentation of residual recipient cells in small samples from bone marrow, peripheral blood and cerebrospinal fluid after bone marrow transplantation.
The long arm of the human Y-chromosome contains about 800 to 5000 copies of the tandemly repeated DNA sequence DYZ1. A major part of the repeating unit (pHY10) has been cloned and sequenced. Primers were designed to match a part of this repeat sequence for the amplification of a 154 bp fragment spanning the EcoRI restriction site of the unit. Typical dilution experiments showed that this polymerase chain reaction (PCR) method allows the detection of 5 to 10 male cells among 100,000 female cells, or in 500 microL of cerebrospinal fluid containing only one cell per microL. In addition, the quality of the DNA used for the amplification reaction is less critical, thus allowing analysis of long-term stored samples such as bone marrow smears or dried blood stains spotted onto filter paper, which might contain partially degraded DNA. We applied this technique to detect residual host cells in the clinical setting of human sex-mismatched bone marrow transplantation (BMT). Fourteen patients, receiving transplantations because of leukemias could be supervised so far. Throughout the whole period of monitoring (days +14 until +911 post BMT; median: 160 days), residual host cells were detected in all but three patients. Persistence of host cells in the early phase post-BMT was mostly transient and probably due to long-term surviving host T-lymphocytes. Reappearance of host cells several months after BMT is highly suspicious of relapse from the underlying malignancy. Due to its high sensitivity, PCR is a valuable tool in monitoring the switch from recipient to donor cell population.(ABSTRACT TRUNCATED AT 250 WORDS)
Suttorp
M
M
Department of Internal Medicine II, University of Kiel, Germany.
Sprenger
C
C
Dreger
P
P
Sievers
E
E
Claviez
A
A
Löffler
H
H
Schaub
J
J
Schmitz
N
N
D001483:Base Sequence; D001773:Blood Cells / Q000473:pathology; D001853:Bone Marrow / Q000473:pathology; D016026:Bone Marrow Transplantation / Q000473:pathology; D002555:Cerebrospinal Fluid / Q000166:cytology; D002678:Chimera; D004247:DNA / Q000235:genetics*; D016172:DNA Fingerprinting / Q000379:methods*; D005260:Female; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000706:statistics & numerical data; D012091:Repetitive Sequences, Nucleic Acid; D012680:Sensitivity and Specificity; D013997:Time Factors; D014019:Tissue Donors; D014998:Y Chromosome
D004247:DNA