S.A. van der Vlies

Maastricht University, Maestricht, Limburg, Netherlands

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Publications (6)12.62 Total impact

  • C.E.M. Voorter · S.A. van der Vlies · E M van den Berg-Loonen
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    ABSTRACT: The large number of polymorphic sites in the HLA-B locus makes sequencing an efficient way of detecting and analysing them. Most polymorphic sites are located in the alpha1 and alpha2 domains of the molecule, encoded by exons 2 and 3 of the gene. An HLA-B-specific sequence-based typing (SBT) strategy was designed for routine application identifying the polymorphic sites in these domains. Exons 2 and 3 were amplified separately using amplification primers located in intron 1, intron 2 and intron 3. Separate amplification of exons 2 and 3 resulted in short polymerase chain reacting (PCR) products and enabled a solid-phase sequencing approach, which made correct assignment of heterozygous positions possible due to low background. A one-step sequencing reaction was performed using fluorescent dye-labelled sequencing primers. One forward sequencing reaction was performed for exon 2, whereas for exon 3, two forward sequencing reactions were needed using two different sequencing primers located in intron 2 and exon 3. The combined sequences of exon 2 and 3 were used for automatic alignment to an HLA-B sequence database and automatic allele assignment. A total of 355 individuals with at least one allele belonging to the B7 cross-reacting group (B7, 13, 22, 27, 40, 41, 42, 47, 48, 81 and 82) were typed for HLA-B by SBT. In the B7 group 48 different alleles were identified, in the non-B7 group a further 59 alleles were sequenced, 9 new alleles were identified. The sequencing strategy described has proven to be reliable and efficient for high-resolution HLA-B typing.
    No preview · Article · Nov 2000 · Tissue Antigens
  • C.E.M. Voorter · S van der Vlies · M Kik · E M van den Berg-Loonen
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    ABSTRACT: The Bw4 and Bw6 epitopes were the first HLA-B differences to be recognized by serological methods. Since then 44 serological groups have been identified and more than 250 alleles assigned by molecular typing methods. In general each serological HLA-B group is associated with the presence of either the Bw4 or the Bw6 epitope. There are several exceptions to this rule. Four alleles, B*4601, *7301, *5503 and *1806, show no serological reactivity with either Bw4 or Bw6. Although the Bw6 motif at residues 77-83 is present in these alleles the Bw6 epitope is modified by a valine at residue 76. One or more alleles from the B8, B40 and B62 groups are identified as Bw4 positive, whereas all others are Bw6 positive. In the groups B27, B44 and B47 several alleles are found to be Bw6 positive, while the majority is Bw4 positive. Histocompatibility testing of dialysis patients and their families revealed the serological presence of an unexpected Bw4 epitope associated with B18 in one patient and B56 in another. Allele-specific amplification and sequencing of exons 2 and 3 of these HLA-B alleles revealed the presence of the Bw4 sequence motif for both. The new alleles were assigned B*1809 and B*5607, respectively. In 2 other patients the presence of a new B*07 allele was determined by sequence based typing. Although the new allele, B*0715, showed the Bw6 sequence motif at positions 77 to 83, a substitution of amino acid 76 from glutamic acid to valine was identified. This change resulted in an aberrant Bw6 serological reaction pattern.
    No preview · Article · Nov 2000 · Tissue Antigens
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    ABSTRACT: Unrelated Bubi, native to the island of Bioko (Equatorial Guinea), were previously typed by low-resolution polymerase chain reaction using sequence-specific primers (PCR-SSP) and serology for HLA-A, -B and -C. HLA-B*44 was found frequently and associated with Cw*07. We have studied the HLA subtypes of 20 B*44pos/Cw*07pos Bubi individuals. HLA-B and -C were typed by sequencing exons 2 and 3. To distinguish the alleles Cw*1701/02/03, Cw*07011/012/06 and Cw*1801/02 additional sequencing of exon 1 or 5 was performed. All 20 B*44pos/Cw*07pos individuals of the Bubi population were typed Cw*0706 positive. Nineteen of them carried the B*44032 allele and one B*4407. In addition, 19 B*44neg/ Cw*07pos Bubi individuals were typed for HLA-C and none of them proved Cw*0706 positive. To determine whether the association between Cw*0706 and B*44032 was limited to the Bubi, 19 individuals from Dutch Caucasian families were typed in which B44 and Cw7 segregated on one haplotype. None of these individuals showed the presence of B*44032 or Cw*0706. The haplotypes found in the Dutch Caucasians were B*4402-Cw*0704, B*44031-Cw*07011 and B*44031-Cw*0702. The present observation indicates a strong association between B*44032 and Cw*0706 in the Bubi population.
    No preview · Article · Feb 2000 · Tissue Antigens
  • S.A. van der Vlies · C.E.M. Voorter · E M van den Berg-Loonen
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    ABSTRACT: HLA-C was shown to be a highly polymorphic gene which can be accurately typed for by sequencing methodologies. Most HLA-C sequence-based typing protocols described so far are based on analysis of sequence data of exons 2 and 3. Nonetheless, exons 1, 4 and 5 also contain nucleotide substitutions which contribute to the polymorphisms of the HLA-C locus. Ten alleles contain polymorphic positions in exons 1, 4 and 5, Cw*0701/06, Cw*1202112, Cw*15051/2, Cw*1701/02, and Cw*1801/02. Here we describe a reliable solid-phase sequence-based typing strategy for sequencing exons 1, 4 and 5, which is an extended protocol of our previous HLA-C study. A panel of 16 individuals, carrying 27 different Cw-alleles, was typed for exons 1, 4 and 5 to check the newly designed primers. No allelic dropout or preferential amplification was noticed in these individuals. The panel was also sequenced in order to check the known polymorphisms present in exons 1, 4 and 5. For exon 5 the sequences of the alleles Cw*0302, *0501 and *07011 did not correspond with the published data. In addition, exons 1, 4 and 5 were sequence-based typing typed in 28, 17 and 59 individuals, respectively. Two new alleles were detected which contain polymorphic positions outside exons 2 and 3, Cw*07012 and Cw*1703. The unknown sequence data of exons 1, 4 and 5 of the alleles Cw*02024, *0308, *1506 and *16041 were elucidated. The described high-resolution sequence-based typing protocol for sequencing exons 1, 4 and 5 will be a valuable tool to study the HLA-C locus for polymorphisms outside exons 2 and 3 and for identification of the presently known HLA-C alleles with polymorphic positions in these exons.
    No preview · Article · Sep 1999 · Tissue Antigens
  • S A van der Vlies · C.E.M. Voorter · E M van den Berg-Loonen
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    ABSTRACT: Serological typing of HLA-C has been poor and almost half of its alleles are serologically undetectable blanks in most populations. Therefore, DNA typing techniques have been used to identify and type for the HLA-C gene. Sequence-based typing (SBT) has proven a major typing strategy for highly polymorphic HLA genes. The technique enables direct identification of all sequence motifs without the need to continuously adjust primers. Here we describe a reliable solid-phase SBT strategy for HLA-C which can be used to distinguish all currently known HLA-C alleles without prior knowledge gained by low resolution typing. Exons 2 and 3 were amplified and sequenced and if necessary sequences of exons 1 and 5 were determined. A total of 257 individuals were typed for HLA-C using this protocol and 30 of the 42 known HLA-C alleles were detected. All heterozygous combinations found in this study were unambiguously discriminated. One hundred and forty-four individuals from the Dutch population were typed randomly. In this group Cw*0701 and *0702 were the most frequently detected alleles. Of the serological Cw blank alleles Cw*1203 was found to have the highest frequency (16%). From the total group 212 individuals were typed serologically and 106 were retyped with 97 selected antisera to further compare serological and molecular defined phenorypes. Discrepancies between serological typing and SBT are mainly attributable to the serologically Cw blank alleles Cw*12–18. The high resolution SBT protocol described will be a valuable tool for the identification of HLA-C alleles and the determination of the role of HLA-C in marrow and organ transplantation.
    No preview · Article · Jan 1999 · Tissue Antigens
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    ABSTRACT: Genomic heterogeneity has been observed in several solid tumor types. To investigate this phenomenon in head and neck squamous cell carcinoma (HNSCC), we analyzed macroscopically distinct tissue samples of 12 resected tumors by a combination of fluorescence in situ hybridization (FISH) and DNA flow cytometry. Using a panel of centromeric DNA probes, numerical chromosomal aberrations were detected in 10 tumors, 9 of which showed a single DNA aneuploid peak. Imbalances in chromosomal copy numbers resulted in unique patterns of chromosomal aberrations for each tumor case. Two types of tumors could be distinguished, i.e., tumors (n = 5) containing a single aneusomic clone and tumors (n = 5) with multiple aneusomic clones. The center of this latter group of tumors was shown to be genetically more heterogeneous than the tumor margin. In conclusion, this study showed that 1) the pattern of chromosomal aberrations varies greatly between different HNSCC, 2) a major clone with a specific pattern of chromosomal aberrations has spread throughout most HNSCC, and 3) a subgroup of HNSCCs contains additional clones with a different pattern of chromosomal aberrations. Based on these results, HNSCC can be divided into a genetically more homogeneous and a genetically more heterogeneous group.
    Full-text · Article · Jun 1998 · Cytometry

Publication Stats

162 Citations
12.62 Total Impact Points

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  • 1998-1999
    • Maastricht University
      • Department of Genetics and Molecular Cell Biology
      Maestricht, Limburg, Netherlands