New Method for DNA Microarrays Development: Applied to Human Platelet Antigens Polymorphisms

Abstract

DNA microarrays are a powerful experimental tool for the detection of specific genomic sequences and are invaluable to a broad array of applications: clinical diagnosis, personalized medicine, drug research and development, gene therapy, food control technologies, and environmental sciences. Alloimmunization to human platelet antigens (HPAs) is commonly responsible for neonatal alloimmune thrombocytopenia, post-transfusional purpura and platelet transfusion refractoriness. Using DNA microarrays, we developed a diagnosis to type the biallelic HPA-1 platelet group. The region for the human genomic DNA sequence that contains the polymorphism responsible for HPA-1 alleles was amplified by polymerase chain reaction (PCR). The expected DNA fragments were hybridized on DNA microarrays, and the data were analyzed using specially developed software. Our initial results show that the two HPA-1 antigens polymorphisms containing a single base difference were detected using DNA microarrays.

High‐throughput simultaneous genotyping of human platelet antigen‐1 to ‐16 by using suspension array

“…HPA‐genotyping is useful for the diagnosis and prevention of the PLT alloimmune syndromes. Many methods for HPA genotyping have been reported in recent years, such as PCR–sequence‐specific primer, PCR–allele‐specific oligonucleotide, PCR‐RFLP, PCR‐SBT, real‐time quantitative PCR, and DNA microarray. While each of these methods has its unique advantages, most of them cannot perform high‐throughput simultaneous testing of all types of HPAs and need to be further improved in terms of specificity, sensitivity, and reliability.…”

“…Many HPA‐genotyping methods have been reported in recent years, such as polymerase chain reaction (PCR)–sequence‐specific primer, PCR–allele‐specific oligonucleotide, PCR–restriction fragment length polymorphism (PCR‐RFLP), PCR–sequence‐based typing (PCR‐SBT), real‐time quantitative PCR, and DNA microarray . Although all these methods involve PCR amplification, they show that differences differ in the design of PCR primers and detection of PCR products.…”

Evanescent wave fluorescence biosensor combined with DNA bio-barcode assay for platelet genotyping

“…Oligonucleotide probes used for the presented BBC assay were first qualified using biochip technology, as described in a previous work (Bres et al, 2005). Sequences corresponding to BBC/cBBC, capture/target and probe/target were proved not to cross-hybridize.…”

“…A first evaluation of our BBC assay/fluorescence biosensor was carried out using 84mers synthetic oligonucleotides as targets. These targets correspond to real sequences routinely used for platelet genotyping (Metcalfe et al, 2003;Bres et al, 2005). After identification of beads biofunctionalization conditions, selectivity and sensitivity of the assay were investigated and compared to literature.…”

Detection of blood‐transmissible agents: can screening be miniaturized?

“…Consequently, in 2001, while considering their potential application in blood testing, Petrik 16 estimated that a limited number of probes (300‐400) would allow the analysis of blood groups, platelet and granulocyte antigens, and the screening of infection. The concept later exploited in immunohematology consisted of the multiplex amplification of genomic DNA and optical detection of the amplification products after their hybridization onto probes immobilized onto chips, 17‐19 384‐well microplates, 20 or beads 21 . While protein chips are proving to be more complex to develop than DNA chips, experts agree on their likely strong impact on immunohematology allowing the detection of certain phenotypic modifications that escape genetic analysis 22‐26 .…”