Figure 1. The FastScan™ β-Amyloid ELISA Kit #44261 detects human Aβ-43, Aβ-42, Aβ-40, Aβ-39, Aβ-38, and Aβ-37, but not pE3 peptide. The absorbance at 450 nm for a panel of human β-amyloid peptides is shown in the upper figure. The corresponding western blot of a panel of human β-amyloid peptides using β-Amyloid antibody is shown in the lower figure.
Figure 2. The relationship between the concentration for each human β-amyloid peptide and the absorbance at 450 nm using the FastScan™ β-Amyloid ELISA Kit #44261 is shown.
|Product Includes||Volume (with Count)||Solution Color|
|FastScan™ ELISA Microwell Strip Plate, 96 Well 53257||1 x 96 tests|
|β-Amyloid Mouse Capture mAb||1 x 1 ea||Green (Lyophilized)|
|β-Amyloid Rabbit HRP-linked mAb||1 x 1 ea||Red (Lyophilized)|
|FastScan™ ELISA Capture Antibody Diluent||1 x 3 ml||Green|
|FastScan™ ELISA HRP Antibody Diluent||1 x 3 ml|
|TMB Substrate 7004||1 x 11 ml|
|STOP Solution 7002||1 x 11 ml|
|Sealing Tape||1 x 1 ea|
|ELISA Wash Buffer (20X) 9801||1 x 25 ml|
|FastScan™ ELISA Cell Extraction Buffer (5X) 69905||1 x 10 ml|
|FastScan™ ELISA Cell Extraction Enhancer Solution (50X) 25243||1 x 1 ml|
|FastScan™ ELISA Kit #44261 Positive Control Type 2||1 x 1 ea|
NOTE: Prepare solutions with deionized/purified water or equivalent.
Prepare only as much reagent as needed on the day of the experiment.
*IMPORTANT: The provided FastScan™ ELISA Cell Extraction Enhancer Solution (50X) may precipitate when stored at 4°C. To dissolve, warm briefly at 37°C and mix gently. The FastScan™ ELISA Cell Extraction Enhancer Solution (50X) can be stored at room temperature to avoid precipitation.
NOTE: The 1X Cell Extraction Buffer contains phosphatase inhibitors. Protease inhibitors should be added to the 1X Cell Extraction Buffer immediately prior to lysing cells. Additional phosphatase inhibitors can also be added (e.g. Protease/Phosphatase Inhibitor Cocktail (100X) #5872, not supplied).
NOTE: A select number of FastScan™ ELISA kits do not contain a positive control, please refer to Product Includes table on the datasheet for a list of included reagents. Should you need support on how to generate a positive control for those kits, contact CST technical support at [email protected].
For adherent cells
For suspension cells
NOTE: Equilibrate all materials and prepared reagents to room temperature prior to running the assay.
*NOTE: Certain FastScan™ ELISA Kits may require additional washes at this step. Any requirements for additional washes will be specifically noted in the product “Description” of the kit’s datasheet.
NOTE: Initial color of positive reaction is blue, which changes to yellow upon addition of STOP Solution.
posted May 2018
revised November 2019
Protocol Id: 1924
The FastScan™ β-Amyloid ELISA Kit is a sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of β-amyloid. To perform the assay, sample is incubated with a capture antibody conjugated with a proprietary tag and a second detection antibody linked to HRP, forming a sandwich with β-amyloid in solution. This entire complex is immobilized to the plate via an anti-tag antibody. The wells are then washed to remove unbound material. TMB is then added. The magnitude of observed signal is proportional to the quantity of β-amyloid.
*Antibodies in kit are custom formulations specific to kit.
The FastScan™ β-Amyloid ELISA Kit detects endogenous levels of β-amyloid, as shown in Figure 1. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.
Amyloid β (Aβ) precursor protein (APP) is a 100-140 kDa transmembrane glycoprotein that exists as several isoforms (1). The amino acid sequence of APP contains the amyloid domain, which can be released by a two-step proteolytic cleavage (1). The extracellular deposition and accumulation of the released Aβ fragments form the main components of amyloid plaques in Alzheimer's disease (1). APP can be phosphorylated at several sites, which may affect the proteolytic processing and secretion of this protein (2-5). Phosphorylation at Thr668 (a position corresponding to the APP695 isoform) by cyclin-dependent kinase is cell-cycle dependent and peaks during G2/M phase (4). APP phosphorylated at Thr668 exists in adult rat brain and correlates with cultured neuronal differentiation (5,6).
Explore pathways + proteins related to this product.