IMMOBILIZED BACTERIOPHAGE USED FOR SPECIFIC DETECTION OF E.COLI USING ELECTROCHEMICAL IMPEDANCE SENSINGAbstract
In the present study, bacteriophage based electrochemical analysis is applied to detect pathogenic E.coli bacteria. Phages are parasites that only infect one specific bacteria, can be useful to identify bacterial contaminants in food, water, etc. Phage has been isolated from various water sources such as waste, stagnant and sewage water by overlay method against host bacteria. Titer of phages was calculated to be 107 pfu/ml using 10-fold dilutions. Morphology and identification of phage was done using Transmission Electron Microscopy (TEM) by staining with Uranyl Acetate. The phage capsid was about 78 nanometers (nm) in diameter with tail of length 527 nm as compared to wild type lambda phage whose head is about 65 nm. Plaque reaction activity was observed within 4- 6 hrs against host bacteria by spot test. It is therefore proposed that the isolated phage is lambda like virus and lytic life cycle. Bacteriophages are immobilized onto platinum (pt) electrode by self-assembled molecular monolayer (SAM) and electrochemical impedance analysis performed for a rapid and specific detection of E.coli cell. Impedance spectra for sensors of different concentrations of E .coli are recorded in phosphate buffered saline (PBS) and impedance were obtained in a frequency range from 100mHz to 100kHz, using a modulation voltage 10mV. Rapid and Specific detection with very low concentration of E.coli (104 cfu/ml). This work proposes that measurements of impedance of cell suspensions can provide an alternative method quantifying bacterial cells that is very simple, rapid, economical and a direct means of detecting a specific bacteria.
Vipin Singh, Vidushi Rawal, Saachi Lakhanpal, Pranay Jain *, Swati Dahiya and C.C. Tripathi
Department of Biotechnology Engineering ,University Institute of Engineering and Technology, Kurukshetra University, Kurukshetra, Haryana, India
17 February, 2015
13 April, 2015
05 June, 2015
01 September, 2015