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Global Journal of Analytical Chemistry 2012, 3: 17
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Research Article
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Application of supramolecular DNA-streptavidin complexes for ultrasensitive detection of several toxins by immuno-PCR
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Dmitry Yu. Ryazantsev, Elena E. Petrova, Natalia A. Kalinina, Tatiana I. Valyakina,
Evgeny V. Grishin, Sergei K. Zavriev
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Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation 117997 Moscow, Ul. Miklukho-Maklaya, 16/10
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Abstract |
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Application of supramolecular complexes of DNA-fragments biotinylated at both ends with streptavidin demonstrated ultrasensitive detection of three microbial toxins (Vibrio cholerae toxin, Staphylococcus aureus toxic shock syndrome toxin and S. aureus enterotoxin A) by real-time immuno-PCR. Bisbiotinylated DNA probes of various types were employed to design DNA-streptavidin complexes, i.e. single-stranded DNA of 60 n., double-stranded DNA of 150, 300, and 750 bp-long at different DNA-streptavidin ratio. The test systems displayed the following sensitivities: for SEA - 1 pg/ml, TSST – 0.1 pg/ml, CHT – 1 pg/ml composing 300, 2000, and 200-fold higher sensitivity, respectively, as compared to the corresponding ELISA tests. Interestingly, a high sensitivity can be reached regardless of a DNA probe size. However, the most precise results were observed with a short single chain probe at the DNA-streptavidin ratio 1/1.
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Keywords |
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Toxin detection; Immuno-PCR; Real-time PCR; Monoclonal antibodies; Staphylococcal enterotoxins; Cholera toxin
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