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Surface Plasmon Resonance Based Label-Free Detection of Salmonella using DNA Self Assembly

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Abstract

Typhoid is re-emerging as a biggest health threat to third world countries. One of the major challenge is false negative diagnosis using existing immunodiagnostic methods due to overlapping symptoms of other infections (like brucellosis, malaria, hepatitis) that mimic this enteric fever (typhoid). Surface plasmon resonance (SPR) based DNA hybridisation biosensor has been fabricated by generating self-assembled monolayer of 5′-thiolated single-stranded DNA (ssDNA) probe onto gold surface. Highly specific DNA probe has been selected from conserved Vi capsular antigen gene of Salmonella enterica serovars typhi. This DNA biosensor has been investigated for label-free real-time monitoring of Salmonella. Interestingly, 0.019 ng mL-1 (2 fM) is the lowest detected concentration in PBS with association (k a) and dissociation (k d) rate constants to be k a (M−1 s−1) = 6.68 × 104 ± 2.3 and k d (s−1) = 5.6 × 10−3 ± 0.01, respectively. This biosensor was successfully demonstrated to distinguish complementary, non-complementary and one-base mismatch sequences and reusability up to 40 hybridisation cycles at room temperature. Successful results were obtained for hybridisation studies of genomic DNA isolated from spiked urine sample. Performance characteristics of this biosensing device suggested further scope to fine-tune such DNA-based assays to be implied for clinical, food and environmental applications.

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Acknowledgments

We are grateful to Prof. Sopory, Vice Chancellor, Prof. Subhasis Ghosh, Director, AIRF, JNU, India, for their constant encouragement and infrastructural support. Thanks are due to Ms. Meenakshi and AIRF team members for their kind support and cooperation. DBT-sponsored projects IYBA-2008 (BT/B1/12/045/2008) and RGYI-2009(BT/PR13127/GBD/27/195/2009), UPE1- capacity buildup fund, DST PURSE  are dully acknowledged.

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Authors and Affiliations

  1. Advanced Instrumentation Research Facility, Jawaharlal Nehru University, New Delhi, 110067, India

    Anu Singh & Kavita Arora

  2. School of Life Sciences, Jaipur National University, Jaipur, 302017, India

    Anu Singh & H. N. Verma

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  1. Anu Singh
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  2. H. N. Verma
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Correspondence to Kavita Arora.

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Singh, A., Verma, H.N. & Arora, K. Surface Plasmon Resonance Based Label-Free Detection of Salmonella using DNA Self Assembly. Appl Biochem Biotechnol 175, 1330–1343 (2015). https://doi.org/10.1007/s12010-014-1319-y

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  • DOI: https://doi.org/10.1007/s12010-014-1319-y

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