Colorimetric Detection of a Bacterial Biomarker Using a Modified Litmus Test

Abstract

Source: Tram, K., et al. Colorimetric Detection of Bacteria Using Litmus Test. J. Vis. Exp. (2016).

This video demonstrates a colorimetric E. coli detection method, involving an E. coli biomarker activating a DNAzyme, leading to cleavage of RNA linkage and detachment of a DNAzyme-urease conjugate. A subsequent colorimetric assay detects the E. coli biomarker through ammonia-induced pH changes.

Protocol

1. Preparing E. coli Cells for Testing For a desired cell suspension, transfer 1 ml of cultured stock to a 1.5 ml microfuge tube. Centrifuge the cells at 6,000 x g for 10 min at 4 °C. Carefully remove the supernatant without disturbing the cell pellet. Add 10 µl of reaction buffer to the cell pellet and resuspend the cells. Sonicate the cell suspension for 5 min. Transfer the cell suspension to an ice box for 5 min. Sonicate the cell s…

Materials

Ethylenediaminetetraacetic acid (EDTA)	VWR AMRESCO	105
Sodium Hydroxide (NaOH) pellets	BIO BASIC CANADA INC	SB6789
Tris-base	VWR AMRESCO	497
Urea	VWR AMRESCO	M123
Xylenecyanol FF	SIGMA-ALDRICH	X-4126
0.04% Phenol red	SIGMA-ALDRICH	P3532
10x T4 polynucleotide kinase reaction buffer	Lucigen	30061-1
E. coli K12 (MG1655)	ATCC	ATCC700926
Magnetic Bead (BioMag)	Bangs Laboratories Inc
Magnetic Seperation Rack	New England BioLabs	S1506S
Microfuge tubes	Sarstedt	72.69
Cell culture incubator	Eppendorf Scientific	M13520000
Branson Ultrasonic cleaner	Branson	N/A