FKHRL1 (Phospho-Ser253) ELISA Kit

Sample

cultured cells

Species Reactivity

Human; Mouse; Rat

Intended Use

The FKHRL1 (Phospho-Ser253) Cell-Based ELISA Kit is a convenient, lysate-free, high throughput and sensitive assay kit that can monitor FKHRL1 protein phosphorylation and expression profile in cells. The kit can be used for measuring the relative amounts of phosphorylated FKHRL1 in cultured cells as well as screening for the effects that various treatments, inhibitors (ie. siRNA or chemicals), or activators have on FKHRL1 phosphorylation.
Please read the provided manual entirely prior to use as suggested experimental protocols may have changed.

Contents of Kit

1. 96-Well Cell Culture Clear-Bottom Microplate: 2 plates
2. 10x TBS: 24 mL
3. Quenching Buffer: 24 mL
4. Blocking Buffer: 50 mL
5. 10x Wash Buffer: 50 mL
6. 100x Anti-FKHRL1 (Phospho-Ser253) Antibody (Rabbit Polyclonal): 60 μL, red
7. 100x Anti-FKHRL1 Antibody (Rabbit Polyclonal): 60 μL, purple
8. 100x Anti-GAPDH Antibody (Mouse Monoclonal): 60 μL, green
9. HRP-Conjugated Anti-Rabbit IgG Antibody: 12 mL, glass
10. HRP-Conjugated Anti-Mouse IgG Antibody: 12 mL, glass
11. Primary Antibody Diluent: 12 mL
12. Ready-to-Use Substrate: 12 mL
13. Stop Solution: 12 mL
14. Crystal Violet Solution: 12 mL
15. SDS Solution: 24 mL
16. Adhesive Plate Seals: 4 seals

Storage

Upon receipt, the kit should be stored at 4°C. The un-opened kit will be stable for up to 6 months from the date of shipment if stored at 4°C. Diluted Anti-FKHRL1 (Phospho-Ser253) Antibody, Anti-FKHRL1 Antibody and diluted Anti-GAPDH Antibody can each be stored at 4°C for up to two weeks. HRP-Conjugated Anti-Rabbit IgG Antibody and HRP-Conjugated Anti-Mouse IgG Antibody will be stable at 4°C for up to six months. The SDS Solution should be stored at room temperature or warmed up to room temperature if stored at 4°C.

Reconstitution And Storage

Note: Please remember to allow all solutions to warm up to room temperature prior to use.
##1x TBS## - 1x TBS is used to wash cells seeded on the plate. 1x TBS can be prepared by adding 1 volume of 10x TBS provided in the kit to 9 volumes of ddH2O.
##Fixing Solution## - This solution is NOT provided. Fixing Solution is used to fix cells after cell culture. It is prepared by adding formaldehyde to 1x TBS with light mixing. The 4% formaldehyde is used for adherent cells and 8% formaldehyde is used for suspension cells and loosely attached cells. 37% formaldehyde can be purchased from Sigma Cat# F-8775.
##Quenching Buffer## - This solution is provided as ready-to-use. Quenching Buffer is used to inactivate the endogenous peroxidase activity of the seeded cells.
##Blocking Buffer## - This solution is provided as ready-to-use. Blocking Buffer is used to block additional binding sites in each well.
##Wash Buffer## - This buffer is provided as a 10x solution. 1x Wash Buffer can be prepared by adding 1 volume of 10x Wash Buffer provided in the kit to 9 volumes of ddH2O.
##100x Anti-FKHRL1 (Phospho-Ser253) Antibody## - This antibody is a rabbit polyclonal antibody. This antibody was tested to be specific for the FKHRL1 protein phosphorylated at Ser253. The supplied antibody is a 100x solution. Make 1:100 dilutions in Primary Antibody Diluent prior to use. The diluted primary antibody can be stored at 4°C for up to two weeks.
##100x Anti-FKHRL1 Antibody## - This antibody is a rabbit polyclonal antibody. This antibody was tested to be specific for the FKHRL1 protein. The supplied antibody is a 100x solution. Make 1:100 dilutions in Primary Antibody Diluent prior to use. The diluted primary antibody can be stored at 4°C for up to two weeks.
##100x Anti-GAPDH Antibody## - This antibody is a mouse monoclonal antibody. This antibody was tested to be specific for GAPDH. The supplied antibody is a 100x solution. Make 1:100 dilutions in Primary Antibody Diluent prior to use. The diluted primary antibody can be stored at 4°C for up to two weeks.
##HRP-Conjugated Anti-Rabbit IgG Antibody## - This solution is provided as ready-to-use. HRP-Conjugated Anti-Rabbit IgG Antibody is used as the secondary antibody to detect the target-bound, primary rabbit antibodies.
##HRP-Conjugated Anti-Mouse IgG Antibody## - This solution is provided as ready-to-use. HRP-Conjugated Anti-Mouse IgG Antibody is used as the secondary antibody to detect the target-bound, primary mouse antibodies.
##Primary Antibody Diluent## - This solution is provided as ready-to-use. Use this solution to dilute the provided antibodies.
##Ready-to-Use Substrate## - This solution is provided as ready-to-use. Ready-to-Use Substrate must be warmed to room temperature before use. Keep away from light as this solution is light-sensitive.
##Stop Solution## - This solution is provided as ready-to-use. Stop Solution must be handled with caution as it contains 2 N Sulfuric Acid (H2SO4) and is corrosive. Wear eye protection and gloves when handling.
##Crystal Violet Solution## - This solution is provided as ready-to-use. Crystal Violet is an intense stain used to stain cell nuclei. Avoid contact with skin and clothing.
##SDS Solution## - This solution is provided as ready-to-use. SDS is used to solubilize the Crystal Violet in preparation for cell staining. Store this solution at room temperature or warm up to room temperature if stored at 4°C.