QUANTITATIVE POLYMERASE CHAIN-REACTION USING A DNA HYBRIDIZATION ASSAY BASED ON SURFACE-ACTIVATED MICROPLATES

被引:13
作者
BERNDT, C [1 ]
BEBENROTH, M [1 ]
OEHLSCHLEGEL, K [1 ]
HIEPE, F [1 ]
SCHOSSLER, W [1 ]
机构
[1] IMTEC IMMUNDIAGNOST GMBH,D-16341 ZEPERNICK,GERMANY
关键词
D O I
10.1006/abio.1995.1150
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
A method for DNA quantification on microplates based on the hybridization between single-stranded target and solid-phase bound capture DNA is presented. Binding of the capture DNA to the microplates was attained by surface activation using organosilanes. Detection of hybridized DNA was performed by an enzyme-linked assay taking advantage of the labeled target DNA. Basic test characteristics are described and application examples are given demonstrating its feasibility for a quantitative PCR (QPCR). The QPCR was carried out by coamplifying an immunoglobuline complementarity determining region 3 (CDR3)-coding plasmid DNA with a synthetic internal standard (IS). IS and plasmid CDR3 both shared primer binding sites and produced length-identical amplicons. The amplified DNA was quantified following differential hybridization with IS- and plasmid-specific capture probes. Based on the product ratios of plasmid to IS, the initial amounts of plasmid DNA were calculated. This way, QPCR was shown to be capable of detecting initial concentration differences of plasmid DNA of about 30%. (C) 1995 Academic Press, Inc.
引用
收藏
页码:252 / 257
页数:6
相关论文
共 25 条
  • [1] BAINS W, 1991, CLIN CHEM, V37, P248
  • [2] BERNDT C, 1994, Patent No. 44168020
  • [3] DETECTION AND QUANTITATION OF NEOPLASTIC-CELLS IN ACUTE LYMPHOBLASTIC-LEUKEMIA, BY USE OF THE POLYMERASE CHAIN-REACTION
    BRISCO, MJ
    CONDON, J
    SYKES, PJ
    NEOH, SH
    MORLEY, AA
    [J]. BRITISH JOURNAL OF HAEMATOLOGY, 1991, 79 (02) : 211 - 217
  • [4] QUANTITATIVE ESTIMATION OF MINOR MESSENGER-RNAS BY CDNA-POLYMERASE CHAIN-REACTION - APPLICATION TO DYSTROPHIN MESSENGER-RNA IN CULTURED MYOGENIC AND BRAIN-CELLS
    CHELLY, J
    MONTARRAS, D
    PINSET, C
    BERWALDNETTER, Y
    KAPLAN, JC
    KAHN, A
    [J]. EUROPEAN JOURNAL OF BIOCHEMISTRY, 1990, 187 (03): : 691 - 698
  • [5] OLIGONUCLEOTIDE GENOTYPING OF HLA POLYMORPHISM ON MICROTITRE PLATES
    CROS, P
    ALLIBERT, P
    MANDRAND, B
    TIERCY, JM
    MACH, B
    [J]. LANCET, 1992, 340 (8824) : 870 - 873
  • [6] DAHLEN P, 1993, CLIN CHEM, V39, P1626
  • [7] IMMUNOENZYMATIC TECHNIQUES APPLIED TO THE SPECIFIC DETECTION OF NUCLEIC-ACIDS - A REVIEW
    GUESDON, JL
    [J]. JOURNAL OF IMMUNOLOGICAL METHODS, 1992, 150 (1-2) : 33 - 49
  • [8] HOLODNIY M, 1992, BIOTECHNIQUES, V2, P17
  • [9] A SIMPLE METHOD OF DETECTING AMPLIFIED DNA WITH IMMOBILIZED PROBES ON MICROTITER WELLS
    KAWAI, S
    MAEKAWAJIRI, S
    YAMANE, A
    [J]. ANALYTICAL BIOCHEMISTRY, 1993, 209 (01) : 63 - 69
  • [10] A SENSITIVE NONISOTOPIC HYBRIDIZATION ASSAY FOR HIV-1 DNA
    KELLER, GH
    HUANG, DP
    MANAK, MM
    [J]. ANALYTICAL BIOCHEMISTRY, 1989, 177 (01) : 27 - 32