(a) After incubation with horseradish peroxidase (HRP) conjugated secondary antibody, wash plates with PBS/0.05% Tween-20.
(b) Add 100 μl of TMB solution/well. Incubate for a specific amount of time or until desired signal is achieved. See product datasheet for recommended incubation times for specific antibody pair.
(c) Add 100 μl of STOP Solution.
(d) Read at Absorbance 450 nm.
STOP Solution comes in solution and is ready to use from the bottle.
Store at 4ºC.
STOP Solution is a proprietary solution used to terminate the peroxidase/TMB reaction for ELISA applications. The TMB substrate reacts with immobilized horseradish peroxidase (HRP) conjugated secondary antibodies to produce a blue solution. Color intensity is an indication of analyte level. After attaining the desired intensity, the reaction is terminated by addition of STOP Solution. Upon addition of STOP Solution the color turns from blue to yellow. Absorbance at 450 nm can be read immediately, and the reaction is stable for one hour.
STOP Solution is used to terminate TMB/peroxidase reaction for ELISA applications and is intended for use in combination with CST's PathScan® Sandwich ELISA Antibody Pairs.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.