Multipurpose high-throughput filtering microarrays (HiFi) for DNA and protein assays.
We are reporting here a low cost colorimetric device for high-throughput multiplexed blood group genotyping and allergy diagnosis, displayed as an automated 96-well microtiter plate format. A porous polymeric membrane sealed at the bottom of each well accounts for the sensor support. For each sensing unit, a 6×6 matrix of specific probes is spotted on the external surface of the membrane resulting in 5 mm(2) microarrays. Thanks to the membrane porosity, reagents dispensed into the well can be eliminated through vacuum soaking. This unusual design drastically reduces the assay background signal. The system was first validated on robust models composed of either two complementary oligonucleotide sequences or one allergen/specific rabbit IgG pair. The quality of both oligonucleotide and protein immobilisation on the membrane substrate was then demonstrated together with the capacity to use the arrayed biomolecules as probes for the quantitative detection of specific targets (respectively complementary oligonucleotide and specific antibody). On the basis of these good results, two multiplex assays were developed for crude biological samples testing, focussing on two human in vitro diagnosis applications: a hybridisation assay for multiplex blood group genotyping and a multiparametric immunoassay for allergy diagnosis. In both cases, the transfer to crude biological samples testing was successful i.e. high signal to noise ratio of the stained membranes, reproducibility and good correlation with results obtained using routine testing procedures.
Le Goff
Gaelle C
GC
Equipe Génie Enzymatique, Membranes Biomimétiques et Assemblages Supramoléculaires, Institut de Chimie et Biochimie Moléculaires et Supramoléculaires, Université Lyon 1-CNRS 5246 ICBMS, Bâtiment CPE-43, bd du 11 novembre 1918, 69622 Villeurbanne, Cedex, France.
Desmet
Cloé
C
Brès
Jean-Charles
JC
Rigal
Dominique
D
Blum
Loïc J
LJ
Marquette
Christophe A
CA
D000485:Allergens; D000818:Animals; D000906:Antibodies / Q000032:analysis; D001363:Awards and Prizes; D001483:Base Sequence; D001789:Blood Group Antigens / Q000235:genetics; D001788:Blood Grouping and Crossmatching / Q000295:instrumentation / Q000379:methods; D003124:Colorimetry / Q000295:instrumentation / Q000379:methods; D004247:DNA / Q000032:analysis* / Q000235:genetics; D005374:Filtration; D005838:Genotype; D006801:Humans; D006967:Hypersensitivity / Q000175:diagnosis; D046228:Microarray Analysis / Q000295:instrumentation* / Q000379:methods; D015345:Oligonucleotide Probes / Q000235:genetics; D020641:Polymorphism, Single Nucleotide; D011506:Proteins / Q000032:analysis*; D011817:Rabbits
D000485:Allergens; D000906:Antibodies; D001789:Blood Group Antigens; D015345:Oligonucleotide Probes; D011506:Proteins; D004247:DNA