In Vitro Selection of a Single-Stranded DNA Molecular Recognition Element against Clostridium difficile Toxin B and Sensitive Detection in Human Fecal Matter
Table 1
Systematic evolution of ligands by exponential enrichments (SELEX) scheme for toxin B-specific MRE selection.
Round
Positive selection
Time
Negative selection
Time
1
Immobilized target (IT)
24 hrs
—
—
2
IT
18 hrs
BSA INT
22 hrs
3
IT
13 hrs
BSA INT
26 hrs
4
IT
7 hrs
Exotoxin A INT
22 hrs
5
IT
3 hrs
Exotoxin A INT
26 hrs
6
IT
30 min
BSA INT
24 hrs
7
IT/competitive elution with 20 g/mL free toxin B
5 min/5 min
IT/competitive elution with 20 g/mL free BSA
5 min/5 min
8
IT/competitive elution with 20 g/mL free toxin B
5 sec/5 sec
IT/competitive elution with 20 g/mL free alpha toxin, 1 hour
5 sec/1 hrs
9
IT/competitive elution with 10 g/mL free toxin B
5 sec/5 sec
IT/competitive elution with 20 g/mL free cholera toxin, 1 hour
5 sec/1 hrs
10
IT/competitive elution with 5 g/mL free toxin B
5 sec/5 sec
IT/competitive elution with 20 g/mL free exotoxin A, 1 hour
5 sec/1 hrs
11
IT/competitive elution with 2.5 g/mL free toxin B
5 sec/5 sec
IT/competitive elution with 20 g/mL free BSA, 24 hrs
5 sec/24 hrs
12
IT/competitive elution with 1 g/mL free toxin B
5 sec/5 sec
—
In vitro selection performed for identifying toxin B-specific MRE. Immobilized target (IT) is toxin B conjugated to magnetic beads. Immobilized negative target (INT) is negative targets conjugated to magnetic beads. BSA is the abbreviation for bovine serum albumin. Times listed are incubation times in hours (hrs), minutes (min), or seconds (sec).
