Nucleic acid sensor for M. tuberculosis detection based on surface plasmon resonance

Abstract

Cysteine modified NH 2 -end peptide nucleic acid (PNA) (24-mer) probe and 5¢-thiol end labeled deoxyribonucleic acid (DNA) probes specific to Mycobacterium tuberculosis have been immobilized onto BK-7 gold coated glass plates for the detection of complementary, one-base mismatch, non-complementary targets and complementary target sequence in genomic DNA of Mycobacterium tuberculosis using a surface plasmon resonance (SPR) technique. The DNA/Au and PNA/Au bio-electrodes have been characterized using contact angle, atomic force microscopy (AFM), electrochemical impedance spectroscopy (EIS) and cyclic voltammetric (CV) techniques, respectively. It is revealed that there is a 252 millidegrees SPR angle change in the case of PNA immobilization and 205 millidegrees for DNA immobilization, indicating increased amount of immobilized PNA molecules. Hybridization studies reveal that there is no binding of the non-complementary target to DNA/Au and PNA/Au electrode. Compared to the DNA/Au bioelectrode, PNA/Au electrode has been found to be more efficient for detection of one-base mismatch sequence. The PNA/Au bioelectrode shows better detection limit (1.0 ng ml -1 ) over the DNA-Au bioelectrode (3.0 ng ml -1 ). The values of the association (k a ) and dissociation rate constant (k d ) for the complementary sequence in case of the PNA/Au bioelectrode have been estimated as 8.5 ¥ 10 4 m -1 s -1 and 3.6 ¥ 10 -3 s -1 , respectively.

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

“…It has been found that upon decreasing the concentration of the complementary target significant change in reflectance angle (m o ) observed from 40 to 2 fM. Figure 6 shows improved detection limit (∼500 times) compared to literature ( Table 2) [39][40][41][42][43][44]. Kinetic calculations have been carried out for complementary target having association constant and dissociation constant as k a (m −1 s −1 )=6.68×10 4 ±2.3 and k d (s −1 )= 5.6×10 −3 ±0.01, respectively.…”

Applications of peptide nucleic acids (PNAs) and locked nucleic acids (LNAs) in biosensor development

“…Kinetic data have been reported for some SPR-based applications, and DNA and PNA probes have been compared from an analytical perspective. For instance, Prabhakar et al [72] quantified the values of the association and dissociation rate constants (K a and K d ) for the DNA complementary sequence for PNA/Au (8.5× 10 4 m −1 s −1 and 3.6×10 −3 s −1 , respectively) and DNA/Au (2.5×10 4 m −1 s −1 and 1.1×10 −3 s −1 ) bioelectrode, thus demonstrating that the results were threefold better when PNA probes were used. Furthermore, no binding with the singlebase mismatched DNA target was observed for the PNAAu bioelectrode.…”

Clinical immunosensing of tuberculosis CFP-10 antigen in urine using interferometric optical fiber array

“…We have also explored to develop clinical diagnostic sensors using surface plasmon resonance (SPR) [12][13][14][15]. These techniques are unique in terms of the target molecule, specificity, and signal-producing procedure, and they are all based on a common highly sensitive optical biosensing platform with a label-free sensing and shared signal amplification mechanism.…”