A novel nucleic acid based bio-assay toward recognition of Haemophilus influenza using bioconjugation and DNA...

Area of Science:

• Biotechnology
• Analytical Chemistry
• Microbiology

Background:

• Haemophilus influenza (H. influenza) is a Gram-negative bacterium.
• H. influenza exhibits resistance to penicillin-family antibiotics due to beta-lactamase production and penicillin-binding protein modification.
• Accurate and sensitive detection methods are crucial for monitoring H. influenza.

Purpose of the Study:

• To develop a novel, highly sensitive DNA-based biosensor for the detection of H. influenza.
• To immobilize a specific thiolated DNA probe onto a gold electrode surface.
• To utilize Square Wave Voltammetry (SWV) for monitoring DNA hybridization and detecting H. influenza.

Main Methods:

• Immobilization of a specific thiolated DNA probe (SH-5'-AAT TTT CCA ACT TTT TCA CCT GCA T-3') onto a gold electrode.
• Detection of target H. influenza cDNA sequence (5'-ATG CAG GTG AAA AAG TTG GAA AAT T-3') using Square Wave Voltammetry (SWV) in toluidine blue (TB) solution.
• Characterization of electrode morphology using Field Scanning Electron Microscopy (FE-SEM).

Main Results:

• The developed DNA biosensor achieved a detection limit as low as 1 ZM for the target H. influenza DNA sequence.
• A linear detection range was established from 1 µM down to 1 ZM.
• The biosensor demonstrated selective differentiation between complementary and mismatched DNA sequences (one, double, and triple base mismatches).

Conclusions:

• A novel and highly sensitive DNA-based biosensor for H. influenza detection has been successfully established.
• The developed method offers excellent selectivity, distinguishing target DNA from sequences with mismatches.
• This approach provides a promising tool for the sensitive monitoring of H. influenza.