Selection of Single-Stranded DNA Molecular Recognition Elements against Exotoxin A Using a Novel Decoy-SELEX Method and Sensitive Detection of Exotoxin A in Human Serum

Table 1

Decoy-SELEX scheme for Exotoxin A MRE selection.

Round	Positive selection (+)	Negative selection (−)
1	Immobilized target (IT) 46 hrs, 50 µL	Immobilization substrate (IS) 18 hrs, 50 µL
2	IT 24 hrs, 50 µL	IS 22 hrs, 50 µL
3	IT 18 hrs, 50 µL	—
4	IT 12 hrs, 50 µL	IS 20 hrs, 50 µL
5	IT 8 hrs, 50 µL	—
6	IT 5.5 hrs, 50 µL	BSA immobilized negative target (INT) 24 hrs, 50 µL
7	IT 1 hrs, 50 µL	—
8	IT 1 hrs, 25 µL	Cholera toxin INT 18 hrs, 50 µL
9	IT 1 hrs, 5 µL	—
10	IT 5 min, 5 µL	BSA INT 24 hrs, 50 µL
11	IT 5 sec, 5 µL	IS 21 hrs, 10 µL
12	IT 5 sec, 5 µL	—
13	IT 5 sec, 5 µL; competitive elution with 2 µg free Exotoxin A, 5 sec	IT 5 sec, 5 µL; competitive elution with 1 mg/mL free BSA, 5 min
14	IT 5 sec, competitive elution with 1 µg free Exotoxin A, 5 sec	—

In vitro selection performed for identifying Exotoxin A specific MRE. Immobilized target (IT) is Exotoxin A bound to magnetic beads. Immobilization substrate (IS) is streptavidin-coated magnetic beads blocked with biotin regent. BSA is the abbreviation for bovine serum albumin. Times listed are incubation times in hours (hrs), minutes (min), or seconds (sec).