High Sensitive IFN gamma ELISA Assay

Rodent Obestatin ELISA Assay

$1,120.00

This Rodent Obestatin ELISA Assay is used for quantitative determination of obestatin in mouse/rat serum samples. It has various advantages, such as highly specific and sensitive quantification, no influences with other body fluids or physiological active substances and unnecessity of sample pretreatment. Mouse/rat obestatin standard of this kit is a highly purified synthetic product (purity: higher than 99%). The Eagle Biosciences Mouse/Rat Obestatin ELISA Assay is for Research Use Only and should not be used for diagnostic procedures.

SKU: YK230 Categories: , , ,

Rodent Obestatin ELISA Assay

The Rodent Obestatin ELISA Assay is For Research Use Only

Size: 1×96 wells
Dynamic Range: 0.082 – 20 ng/mL
Incubation Time: 22 hours
Sample Type: Mouse or Rat Serum
Sample Size: 25 μL
Controls Included

Assay Background

Obestatin is a 23 amino acid residues peptide isolated from the rat stomach. The peptide shares the precursor with a food intake stimulating peptide, ghrelin, but possesses reducing effects on food intake, gut motility and body weight. With the use of an antiserum directed against the mouse/rat obestatin, obestatin immunoreactivity (irOBS) was detected in cells of the gastric mucosa and myenteric plexus and in Leydig cells of the testis in Sprague–Dawley rats. Double labeling of myenteric plexus with antisera against obestatin and choline acetyltransferase (ChAT) revealed that nearly all irOBS neurons were ChAT positive and vice versa.
Obestatin (100nM) added to dissociated and cultured rat cerebral cortical neurons elevated cytosolic calcium concentrations [Ca+2]i in a population of cortical neurons. Intracerebroventricular administration of obestatin inhibited water drinking in ad libitum fed and watered rats, and in food and water deprived animals. In addition, obestatin inhibited angiotensin II-induced water drinking in animals provided free access to water and food. Obestatin peptides had no effect on insulin sensitivity as revealed by hypoglycaemic response when co-administered with insulin, supporting a role of obestatin in regulating metabolism through changes of appetite, but indicating no direct actions on glucose homeostasis or insulin secretion. It is supposed that in rats the effects of obestatin on food intake may be secondary to an action of the peptide to inhibit water drinking.
The obestatin concerning study for energy homeostasis and body weight regulation could be expected to have a large development in the future. The mouse/rat obestatin EIA assay kit developed by our laboratory can be used for direct determination of serum obestatin level’s variations and will be a useful tool for further development of obestatin research.

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Additional Information

Assay Principle

This Mouse/Rat Obestatin ELISA Assay kit is for the determination of obestatin in mouse/rat serum samples is based on a competitive enzyme immunoassay using the combination of highly specific antibody to mouse/ rat obestatin and biotin–avidin affinity system. The 96 wells plate is coated with goat anti rabbit IgG, to which biotinylated mouse/rat obestatin, mouse/rat obestatin standard or samples and rabbit anti mouse/rat obestatin antibody are added for competitive immunoreaction. After incubation and plate washing, horse radish peroxidase (HRP) labeled streptavidin (SA) is added, so that HRP labeled SA-biotinylated mouse/rat obestatin-antibody complex is formed on the surface of the wells. Finally, HRP enzyme activity is determined by 3,3’,5,5’-Tetramethylbenzidine (TMB) and the concentration of mouse/rat obestatin is calculated.

Assay Procedure

  1. Before starting assay, bring all the reagents except samples to room temperature (20-30°C).
  2. Add 350µL of washing solution to each well and keep it for about 30 seconds, then aspirate the washing solution in the wells. Repeat this washing procedure further twice (total 3 times). Finally, invert the plate and tap it onto an absorbent surface, such as paper toweling, to ensure blotting free of most residual washing solution.
  3. Fill 50µL of labeled antigen solution into each well first, then introduce 25µL of each of standard solutions (0, 0.082, 0.247, 0.741, 2.222, 6.667, 20ng/mL) or samples and finally add 50µL of mouse/rat obestatin antibody into each well.
  4. Cover the plate with Adhesive Foil and incubate it at 4°C for 18 – 20 hours (still, no shaking).
  5. After 4°C incubation, take off the Adhesive Foil, aspirate and wash the wells three times as step 2 with approximately 0.35mL/well of washing solution each time.
  6. Pipette 100µL of SA-HRP Solution into each well.
  7. Cover the plate with Adhesive Foil and incubate it at room temperature for 1 hour (still, no shaking).
  8. Take off the Adhesive Foil, aspirate and wash the wells five times as step 2 with approximately 0.35mL/well of washing solution each time.
  9. Add 100µL of TMB Substrate into each well, cover the plate with Adhesive Foil and keep it for 30 minutes at room temperature under a light-proof condition (still, no shaking).
  10. Add 100µL of Reaction Stopping Solution into each well to stop color reaction.
  11. Read the optical absorbance of the wells at 450nm.
  12. Calculate mean absorbance values of standards and plot a standard curve on semi-logarithmic graph paper (abscissa: concentration of standard; ordinate: absorbance values).
  13. Use the standard curve to read mouse/rat obestatin concentrations in samples from the corresponding absorbance values.

Typical Standard Curve

Manual

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