Surface plasmon resonance based label-free detection of Salmonella using DNA self assembly.

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 × 10(4) ± 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.

Monoclonal antibody to single-stranded DNA: a potential tool for DNA repair studies.

Growing evidence suggests that DNA repair capacity is an important factor in cancer risk and is therefore essential to assess. Immunochemical assays are amenable to the detection of repair products in complex matrices, such as urine, facilitating noninvasive measurements, although diet and extra-DNA sources of lesion can confound interpretation. The production of single-stranded, lesion-containing DNA oligomers characterises nucleotide excision repair (NER) and hence defines the repair pathway from which a lesion may be derived. Herein we describe the characterisation of a monoclonal antibody which recognises guanine moieties in single-stranded DNA. Application of this antibody in ELISA, demonstrated such oligomers in supernatants from repair-proficient cells post-insult. Testing of urine samples from volunteers demonstrated a relationship between oligomer levels and two urinary DNA damage products, thymine dimers and 8-oxo-2'-deoxyguanosine, supporting our hypothesis that NER gives rise to lesion-containing oligomers which are specific targets for the investigation of DNA repair.