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

• Published in: Applied biochemistry and biotechnology 2015-02, Vol.175 (3), p.1330-1343
• Main Authors: Singh, Anu, Verma, H. N, Arora, Kavita
• Format: Article
• Language: English
• Subjects: antigens; Biochemistry; Biosensors; Biotechnology; brucellosis; Chemistry; Chemistry and Materials Science; Deoxyribonucleic acid; developing countries; Diagnostic tests; dissociation; DNA; DNA, Single-Stranded - metabolism; DNA, Single-Stranded - urine; fever; genes; genomics; gold; Gold - chemistry; Health risks; hepatitis; Humans; hybridization; Immobilized Nucleic Acids - metabolism; Malaria; Microscopy, Fluorescence; monitoring; Nucleic Acid Hybridization; Recycling; Reproducibility of Results; Resonance; Salmonella; Salmonella enterica subsp. enterica serovar Typhi; Salmonella typhi - isolation & purification; single-stranded DNA; Spectroscopy, Fourier Transform Infrared; Staining and Labeling; Sulfhydryl Compounds - metabolism; surface plasmon resonance; Surface Plasmon Resonance - methods; Typhoid; typhoid fever; urine; Vector-borne diseases
• ISSN: 0273-2289; 1559-0291
• DOI: 10.1007/s12010-014-1319-y

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 (kₐ) and dissociation (kd) rate constants to be kₐ(M⁻¹ s⁻¹) = 6.68 × 10⁴ ± 2.3 and kd(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.