A novel DNA based bioassay toward ultrasensitive detection of Brucella using gold nanoparticles supported histidine: A new platform for the assay of bacteria in the cultured and human biofluids with and without polymerase chain reactions (PCR)

Abstract

Brucella organisms, which are small aerobic intracellular coccobacilli, localize in the reproductive organs of host animals, causing abortions and sterility. In this work, we used a novel method to preparation of excellent genosensor on the surface of low-toxic substrate (gold nanoparticles) supported histidine prepared by fully electrodeposition method. The results of the present work show that the nano-Au-Hist provide suitable active sites for the DNA probe immobilization. The fabricated DNA genosensor employs cyclic voltammetry (CV), and square wave voltammetry (SWV) techniques for monitoring the behavior of the redox probe. To survey the morphological pattern and surface structural characterizations, the Field Emission-Scanning Electron Microscopy (FE-SEM) has been applied. In summary, the gold nanoparticles supported by histidine was checked for immobilization of a Brucella-specific probe and detection of hybridization with a variety of sequences with a high sensitivity. The high sensitivity would be related to more favorable conformation and deflection angle of the probe for an efficient hybridization, higher surface concentration of the probe, and/or enhanced diffusion regime. These lead to better display of the entangled target sequences arising from the nanobiotechnology. The proposed genosensor showed a perfect distinction between complementary, non-complementary and mismatched DNA sequences. The engineered genosensor for detection of the complementary/non-complementary sequences were assayed. The fabricated genosensor was evaluated for the assay of the bacteria in the cultured and human samples with and without polymerase chain reactions (PCR). The genosensor could detect the complementary sequence in linear concentration range of 1 × 10−1 to 1 × 10−10 μM, and a low limit of quantification 0.1 pM.