Rat CRF / CRH EIA

Cell Culture Supernatants
Serum

Colorimetric

96-well Microplate

Pre-
Coated 96-well Strip Microplate
Wash Buffer
Standard Peptide
Assay Diluent(s)
Biotinylated Peptide
H
R
P-
Streptavidin
T
M
B One-
Step Substrate
Stop Solution
Assay Diagram
Positive Control Sample
Capture Antibody
User Manual

Prepare all reagents, samples and standards as instructed.
Add 100 µl detection antibody to each well.
Incubate 1.5 h at R
T or O/
N at 4°
C.
Add 100 µl standard or sample to each well.
Incubate 2.5 h at R
T.
Add 100 µl prepared streptavidin solution.
Incubate 45 min at R
T.
Add 100 µl T
M
B One-
Step Substrate Reagent to each well.
Incubate 30 min at R
T.
Add 50 µl Stop Solution to each well.
Read plate at 450 nm immediately.

Corticotropin-releasing hormone (CRH), corticotropin-releasing factor (CRF), corticoliberin

Corticotropin-releasing factor (CRF) (also known as Corticotropin-releasing hormone (CRH) is a 41-amino acid peptide hormone secreted by the paraventricular nucleus (PVN) of the hypothalamus in response to stress. Its main function is the stimulation of the pituitary synthesis of adrenocorticotropic hormone (ACTH), as part of the hypothalamic–pituitary–adrenal axis (HPA axis). CRF levels may be associated with Alzheimer's disease and major depression. CRF is also synthesized in peripheral tissues, such as T lymphocytes, and is highly expressed in the placenta, suggesting roles in parturition. The RayBiotech CRF Enzyme Immunoassay (EIA) Kit is an in vitro quantitative assay for detecting CRF peptide based on the competitive enzyme immunoassay principle. In this assay, a biotinylated CRF peptide is spiked into the samples and standards. The samples and standards are then added to the plate, where the biotinylated CRF peptide competes with endogenous (unlabeled) CRF for binding to the anti-CRF antibody. After a wash step, any bound biotinylated CRF then interacts with horseradish peroxidase (HRP)-streptavidin, which catalyzes a color development reaction. The intensity of the colorimetric signal is directly proportional to the amount of captured biotinylated CRF peptide and inversely proportional to the amount of endogenous CRF in the standard or samples. A standard curve of known concentration of CRF peptide can be established and the concentration of CRF peptide in the samples can be calculated accordingly.