useful for quantitative determination of human CRF in plasma

Human CRF ELISA kit


print

Background

Corticotropin releasing factor (CRF, also CRH) was initially isolated from ovine hypothalamus by Vale et al., in 1981, and identified as a novel neuropeptide comprising 41 amino acid residues with molecular weight 4758 1). Later human CRF2) and rat CRF3) were also isolated and identified. The mouse CRF peptide is identical at amino acid level to the rat and human CRF peptides4). CRF in anterior pituitary promotes the synthesis and secretion of ACTH, a main factor of hypothalamus-pituitary-adrenal (HPA) axis. In the rat and human, CRF distributes mainly in hypothalamus, but it was also found in spinal cord, stomach, spleen, duodenum, adrenal and placenta. In addition, immunochemical evidence supported the wide distribution of the peptide throughout the central nervous system (CNS) such as olfactory bulb, retina and central auditory system in the rat.

In mouse brain extracts, the highest concentrations of CRF-like immunoreactivity (CRF-LI) has been detected in median eminence and hypothalamus and also existing in amygdala, thalamus, frontal cortex, medulla/pons and cerebellum by radioimmunoassay5). However because of the wide distribution, it is still disputing about CRF whether its blood level can reflect only the function of HPA axis 6).

The relationships between CRF and stress, CRF and Alzheimer disease (AD) were attracted much attention recently. In fact the peptide was also suggested to regulate endocrine, autonomic and behavioral responses to stress, based on an experiment with acute and chronic stress rat models that showed endocrine function changes similar to those seen in patients with depression 6j CRF in serial cerebrospinal fluidiCSFjof patients with depression was strikingly reduced as compared to those of normal subjects 7) , 8j. The mean CRF and ACTH levels in the CSF of AD patients were significantly lower than those of healthy controls 9).  Only in the cortices of those with mild dementia, CRF was reduced significantly. Thus CRF was proposed to serve as a potential neurochemical marker of early dementia and possibly early AD 10).

A large proportion of CRF in human brain was shown to be in the form of complex with its binding protein (CRF-BP). CRF molecule in the complex is unavailable for activation of the CRF receptor. Accordingly reduction in total CRF do not necessarily predict reduction of bioactive free CRF, and the levels of total CRF and CRF in the form of complex (CRF/CRF-BP) were suggested to be the main factors determining the quantity of bioactive free CRF in human brain 11). In AD there have been observed dramatic reduction in the content of free CRF in brain and thus displacement of CRF from CRF-BP was proposed as a possible treatment for AD 12j. In primary neuron culture, CRF exhibited protective effect against cell death induced by amyloid-beta peptide, suggesting that disturbances in HPA axis function can occur independently of alteration in CRF mRNA levels in AD brain and further suggesting an additional role for CRF in protecting neurons against cell death 13). On the other hand, Yanaihara et al. demonstrated immunoreative CRF in various neuroendocrine tumors, and suggested that the blood level of the peptide might be used as a tumor marker 14j

Reference

Feature and Advantages

This ELISA kit is used for quantitative determination of human CRF in plasma (Pretreatment of plasma before assay is necessary). The kit is characterized by its sensitive quantification and high specificity. In addition, it has no influence by other components in samples. CRF standard is highly purified synthetic product.

• The assay kit can measure human CRF within the range of 0.078-2.5 ng/mL.
• The assay is completed within 7.5 hr.
• With one assay kit, 41 samples can be measured in duplicate.
• Test sample: Human plasma (Pretreatment of plasma before assay is necessary)
                     Sample volume: 50 μL
• The 96-wells plate in kit is consisted by 8-wells strips, and the strips can be used separately.
• Precision and reproducibility
      Intra-assay CVi“j  Human plasma 1.17-4.34
      Inter-assay CVi“j  Human plasma 1.46-9.15
• Stability and Storage
      Store all of the components at 2-8Ž.
      The kit is stable under the condition for 6 months from the date of manufacturing.
      The expiry date is stated on the package.

< Specificity>
This ELISA kit has high specificity to CRF, and shows no crossreactivity to ACTH, urocortin 1, urocortin 2 (mouse) and urocortin 3 (mouse, rat).

< Assay Principle>
This ELISA kit for determination of human CRF is based on a sandwich enzyme immunoassay. To the wells of plate coated with highly purified antibody against CRF, standards or samples are added for the 1st step immunoreaction. After the 1st step incubation and plate washing, labeled antibody solution (biotinylated rabbit anti CRF antibody) is added as the 2nd step to form antibody - antigen - labeled antibody complex on the surface of the wells. After the 2nd step incubation and rinsing out excess labeled antibody, horseradish peroxidase (HRP) labeled streptoavidin (SA) is added for binding to labeled antibody. Finally, HRP enzyme activity is determined by 3,3’,5,5’-Tetramethylbenzidine (TMB) and the concentration of human CRF is calculated.

Composition

Component Form Quantity Main@Ingredient
1. Antibody coated plate Microtiter plate 1 plate(96 wells) Rabbit anti mouse/rat CRF antibody
2. Standard antigen Lyophilized 1 vial (2.5 ng) Synthetic mouse/rat CRF(1-41)
3. Labeled antibody solution Liquid 1 bottle (12 mL) Biotinylated rabbit a anti CRF antibody
4. SA-HRP solution Liquid 1 bottle (12 mL) HRP labeled streptoavidin
5. Enzyme substrate solution LLiquid 1 bottle (12 mL) 3,3’,5,5’-tetramethyl benzidine (TMB)
6. Reaction stopping solution liquid 1 bottle (12 mL) 1M  H2SO4
7. Buffer solution liquid 1 bottle (20 mL) Buffer containing a reaction accelerator
8. Washing solution
(concentrated)
liquid 1 bottle (50 mL) Concentrated saline
9. Adhesive sheet   4 pieces Concentrated saline
Product List

Product Name Cat# Quantity Price

CRF ELISA, Human DataSheet

YII-YK132-EX

1 KIT ¥79,000
$1054

Product Name Cat# Quantity Price

CRF, ELISA (High Sensitivity) DataSheet ,

YII-YK131-EX

1 KIT ¥79,000
$1054

Related Products [ antibody ]


Product Name Cat# Quantity Price

Anti CRF (24-41), Human (Rabbit) Unlabeled DataSheet

YII-Y210-EX

50 UL ¥35,000
$467

Antigen F CRF (24-41)(Human, Mouse, Rat)
IHC F 1:1,000-5,000
RIA F 1:35,000
Specificity F CRF (24-41) (human, mouse, rat) 100%, CRF (human, mouse, rat) ~ 100%


Product Name Cat# Quantity Price

Anti CRF (3-41), Human (Rabbit) Unlabeled DataSheet

YII-Y211-EX

50 UL ¥35,000
$467

Antigen F CRF (3-41)(Human, Mouse, Rat)
IHC F 1:1,000-5,000
RIA F 1:7,000
Specificity F Based on ELISA : CRF (3-41) (human, mouse, rat) 100%, CRF (human, mouse, rat) 100%,
                 CRF (17-41) (human, mouse, rat ) < 0.1%, urocortin (human) < 0.01%
                 urocortin (rat) < 0.01%, ACTH (human) < 0.01%, ACTH (mouse, rat) < 0.01%

To be used for research only. DO NOT use for human gene therapy or clinical diagnosis.



“Human CRF ELISA kit” also belongs to the following categories.