ELISA Kit for Orexin A (OXA) Homo sapiens (Human) Competition ELISA

Hypocretin-1

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  • ELISA Kit for Orexin A (OXA) Packages (Simulation)
  • ELISA Kit for Orexin A (OXA) Packages (Simulation)
  • ELISA Kit for Orexin A (OXA) Results demonstration
  • CEA607Hu.jpg Typical Standard Curve
  • Certificate ISO9001: 2008, ISO13485: 2003 Registered

Recovery

Matrices listed below were spiked with certain level of recombinant Orexin A (OXA) and the recovery rates were calculated by comparing the measured value to the expected amount of Orexin A (OXA) in samples.

Matrix Recovery range (%) Average(%)
serum(n=5) 87-101 92
EDTA plasma(n=5) 84-91 88
heparin plasma(n=5) 98-105 102

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Orexin A (OXA) were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level Orexin A (OXA) were tested on 3 different plates, 8 replicates in each plate.
CV(%) = SD/meanX100
Intra-Assay: CV<10%
Inter-Assay: CV<12%

Linearity

The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Orexin A (OXA) and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.

Sample 1:2 1:4 1:8 1:16
serum(n=5) 89-101% 88-101% 94-103% 82-103%
EDTA plasma(n=5) 85-95% 79-91% 83-97% 89-104%
heparin plasma(n=5) 78-89% 92-101% 92-105% 89-103%

Stability

The stability of kit is determined by the loss rate of activity. The loss rate of this kit is less than 5% within the expiration date under appropriate storage condition.
To minimize extra influence on the performance, operation procedures and lab conditions, especially room temperature, air humidity, incubator temperature should be strictly controlled. It is also strongly suggested that the whole assay is performed by the same operator from the beginning to the end.

Reagents and materials provided

Reagents Quantity Reagents Quantity
Pre-coated, ready to use 96-well strip plate 1 Plate sealer for 96 wells 4
Standard 2 Standard Diluent 1×20mL
Detection Reagent A 1 Assay Diluent A 1×12mL
Detection Reagent B 1×120µL Assay Diluent B 1×12mL
Reagent Diluent 1×300µL Stop Solution 1×6mL
TMB Substrate 1×9mL Instruction manual 1
Wash Buffer (30 × concentrate) 1×20mL

Assay procedure summary

1. Prepare all reagents, samples and standards;
2. Add 50µL standard or sample to each well.
    And then add 50µL prepared Detection Reagent A immediately.
    Shake and mix. Incubate 1 hour at 37°C;
3. Aspirate and wash 3 times;
4. Add 100µL prepared Detection Reagent B. Incubate 30 minutes at 37°C;
5. Aspirate and wash 5 times;
6. Add 90µL Substrate Solution. Incubate 10-20 minutes at 37°C;
7. Add 50µL Stop Solution. Read at 450 nm immediately.

ELISA Kit for Orexin A (OXA)

Test principle

This assay employs the competitive inhibition enzyme immunoassay technique. A monoclonal antibody specific to Orexin A (OXA) has been pre-coated onto a microplate. A competitive inhibition reaction is launched between biotin labeled Orexin A (OXA) and unlabeled Orexin A (OXA) (Standards or samples) with the pre-coated antibody specific to Orexin A (OXA). After incubation the unbound conjugate is washed off. Next, avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. The amount of bound HRP conjugate is reverse proportional to the concentration of Orexin A (OXA) in the sample. After addition of the substrate solution, the intensity of color developed is reverse proportional to the concentration of Orexin A (OXA) in the sample.

Citations

  • Neuro-Glial and Systemic Mechanisms of Pathological Responses in Rat Models of Primary Blast Overpressure Compared to “Composite” BlastPubMed: PMC3275793
  • Pentylenetetrazol-induced seizures are exacerbated by sleep deprivation through orexin receptor-mediated hippocampal cell proliferationPubmed: 23852314
  • Orexin-A-induced ERK1/2 activation reverses impaired spatial learning and memory in pentylenetetrazol-kindled rats via OX1R-mediated hippocampal neurogenesis.Pubmed: 24321199
  • Orexin in Patients with Alcohol Dependence Treated for Relapse Prevention: A Pilot StudyPubMed: 26597795
  • Evaluation of Antidiabetic And Insulintropic Potential of Nigella Sativa Seeds Water Extract and /or Alpha Lipoic Acid in Type 2 Diabetic RatsIssues%20Pdf: Vol.2
  • Correlation of Brain Neuropeptide (Nesfatin-1 and Orexin-A) Concentrations with Anthropometric and Biochemical Parameters in Malnourished ChildrenPubMed: 26831553
  • Resuscitation therapy for traumatic brain injury-induced coma in rats: mechanisms of median nerve electrical stimulationPubMed: 26170820
  • Wake-promoting actions of median nerve stimulation in TBI-induced coma: An investigation of orexin-A and orexin receptor 1 in the hypothalamic regionPubMed: 26059340
  • Application of the Co-culture Membrane System Pointed to a Protective Role of Catestatin on Hippocampal Plus Hypothalamic Neurons Exposed to Oxygen and Glucose Deprivationarticle:10.1007
  • Wake-promoting effects of vagus nerve stimulation after traumatic brain injury: upregulation of orexin-A and orexin receptor type 1 expression in the prefrontal …Pubmed:29557373
  • Recurrent circadian fasting (RCF) improves blood pressure, biomarkers of cardiometabolic risk and regulates inflammation in menPubmed: 31426866

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