Orexin A ELISA Kit

Produktnummer ABIN6958316

Produktdetails Orexin A ELISA Kit

Target: Orexin A (OXA)

Reaktivität: Ratte

Nachweismethode: Colorimetric

Methodentyp: Competition ELISA

Detektionsbereich: 12.35 pg/mL - 1000 pg/mL

Untere Nachweisgrenze: 12.35 pg/mL

Applikation: ELISA

Verwendungszweck: The kit is a competitive inhibition enzyme immunoassay technique for the in vitro quantitative measurement of OXA in rat serum, plasma, tissue homogenates, cell lysates, cell culture supernates.

Proben: Cell Culture Supernatant, Cell Lysate, Plasma, Serum, Tissue Homogenate

Analytische Methode: Quantitative

Spezifität: This assay has high sensitivity and excellent specificity for detection of Orexin A (OXA)

Sensitivität: 5.07 pg/mL

Bestandteile:

• Pre-coated, ready to use 96-well strip plate, flat buttom
• Plate sealer for 96 wells
• Reference Standard
• Standard Diluent
• Detection Reagent A
• Detection Reagent B
• Assay Diluent A
• Assay Diluent B
• Reagent Diluent (if Detection Reagent is lyophilized)
• TMB Substrate
• Stop Solution
• Wash Buffer (30 x concentrate)
• Instruction manual

Anwendungsinformationen

Kommentare:

Information on standard material:
The standard might be recombinant protein or natural protein, that will depend on the specific kit. Moreover, the expression system is E.coli or yeast or mammal cell. There is 0.05% proclin 300 in the standard as preservative.

Information on reagents:
The stop solution used in the kit is sulfuric acid with concentration of 1 mol/L. And the wash solution is TBS. The standard diluent contains 0.02 % sodium azide, assay diluent A and assay diluent B contain 0.01% sodium azide. Some kits can contain is BSA in them.

Information on antibodies:
The provided antibodies and their host vary in different kits.

Probenmenge: 50 μL

Testdauer: 2 h

Plattentyp: Pre-coated

Protokoll:

1. Prepare all reagents, samples and standards,
2. Add 50μL standard or sample to each well.
   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.

Aufbereitung der Reagenzien:

1. Bring all kit components and samples to room temperature (18-25 °C) before use.
2. Standard - Reconstitute the Standard with 1.0 mL of Standard Diluent, kept for 10 minutes at room temperature, shake gently(not to foam). The concentration of the standard in the stock solution is 1,000pg/mL. Please prepare 5 tubes containing 0.6 mL Standard Diluent and produce a triple dilution series. Mix each tube thoroughly before the next transfer. Set up 5 points of diluted standard such as 1,000pg/mL, 333.33pg/mL, 111.11pg/mL, 37.04pg/mL, 12.35pg/mL, and the last EP tubes with Standard Diluent is the blank as 0pg/mL.
3. Detection Reagent A and Detection Reagent B - Briefly spin or centrifuge the stock Detection A and Detection B before use. Dilute to the working concentration with Assay Diluent A and B, respectively (1:100).
4. Wash Solution - Dilute 20 mL of Wash Solution concentrate (30x) with 580 mL of deionized or distilled water to prepare 600 mL of Wash Solution (1x).
5. TMB substrate - Aspirate the needed dosage of the solution with sterilized tips and do not dump the residual solution into the vial again.

Note:

1. Making serial dilution in the wells directly is not permitted.
2. Prepare standard within 15 minutes before assay. Please do not dissolve the reagents at 37 °C directly.
3. Detection Reagent A and B are sticky solutions, therefore, slowly pipette them to reduce the volume errors.
4. Please carefully reconstitute Standards or working Detection Reagent A and B according to the instruction, and avoid foaming and mix gently until the crystals are completely dissolved. To minimize imprecision caused by pipetting, use small volumes and ensure that pipettors are calibrated. It is recommended to suck more than 10μL for once pipetting.
5. The reconstituted Standards, Detection Reagent A and Detection Reagent B can be used only once.
6. If crystals have formed in the Wash Solution concentrate (30x), warm to room temperature and mix gently until the crystals are completely dissolved.
7. Contaminated water or container for reagent preparation will influence the detection result.

Aufbereitung der Proben:

• It is recommended to use fresh samples without long storage, otherwise protein degradation and denaturation may occur in these samples, leading to false results. Samples should therefore be stored for a short period at 2 - 8 °C or aliquoted at -20 °C (≤1 month) or -80 °C (≤ 3 months). Repeated freeze-thaw cycles should be avoided. Prior to assay, the frozen samples should be slowly thawed and centrifuged to remove precipitates.
• If the sample type is not specified in the instructions, a preliminary test is necessary to determine compatibility with the kit.
• If a lysis buffer is used to prepare tissue homogenates or cell culture supernatant, there is a possibility of causing a deviation due to the introduced chemical substance. The recommended dilution factor is for reference only.
• Please estimate the concentration of the samples before performing the test. If the values are not in the range of the standard curve, the optimal sample dilution for the particular experiment has to be determined.

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

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Vorsichtsmaßnahmen: The Stop Solution suggested for use with this kit is an acid solution. Wear eye, hand, face, and clothing protection when using this material.

Lagerung: 4 °C/-20 °C

Informationen zur Lagerung:

1. For unopened kit: All reagents should be stored according to the labels on the vials. The Standard, Detection Reagent A, Detection Reagent B, and 96-well Strip Plate should be stored at -20 °C upon receipt, while the other reagents should be stored at 4 °C.
2. For opened kits: the remaining reagents must be stored according to the above storage conditions. In addition, please return the unused wells to the foil pouch containing the desiccant and seal the foil pouch with the zipper.

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Haltbarkeit: 6 months

Referenzen für Orexin A ELISA Kit (ABIN6958316)

Dong, Feng: "Wake-promoting effects of vagus nerve stimulation after traumatic brain injury: upregulation of orexin-A and orexin receptor type 1 expression in the prefrontal cortex." in: Neural regeneration research, Vol. 13, Issue 2, pp. 244-251, (2018) (PubMed).

Zhong, Feng, Wang, Wei: "Wake-promoting actions of median nerve stimulation in TBI-induced coma: An investigation of orexin-A and orexin receptor 1 in the hypothalamic region." in: Molecular medicine reports, Vol. 12, Issue 3, pp. 4441-7, (2015) (PubMed).

Feng, Zhong, Wang, Wei: "Resuscitation therapy for traumatic brain injury-induced coma in rats: mechanisms of median nerve electrical stimulation." in: Neural regeneration research, Vol. 10, Issue 4, pp. 594-8, (2015) (PubMed).

Zhao, Zhang, Tang, Ren, Yang, Liu, Tang: "Orexin-A-induced ERK1/2 activation reverses impaired spatial learning and memory in pentylenetetrazol-kindled rats via OX1R-mediated hippocampal neurogenesis." in: Peptides, Vol. 54, pp. 140-7, (2014) (PubMed).

Ni, Zhu, Song, Liu, Tang: "Pentylenetetrazol-induced seizures are exacerbated by sleep deprivation through orexin receptor-mediated hippocampal cell proliferation." in: Neurological sciences : official journal of the Italian Neurological Society and of the Italian Society of Clinical Neurophysiology, Vol. 35, Issue 2, pp. 245-52, (2014) (PubMed).

Svetlov, Prima, Glushakova, Svetlov, Kirk, Gutierrez, Serebruany, Curley, Wang, Hayes: "Neuro-glial and systemic mechanisms of pathological responses in rat models of primary blast overpressure compared to "composite" blast." in: Frontiers in neurology, Vol. 3, pp. 15, (2012) (PubMed).

Plavcan, McWilliams: "Toothpick obstruction of the ureter." in: The Journal of urology, Vol. 139, Issue 1, pp. 114-5, (1988) (PubMed).

Details zu Orexin A

Target: Orexin A (OXA)

Abstract: OXA Produkte

Synonyme: PPOX ELISA Kit, orexin-A ELISA Kit, hypocretin ELISA Kit, hypocretin neuropeptide precursor ELISA Kit, Hcrt ELISA Kit, HCRT ELISA Kit

Pathways: Synaptic Membrane, Feeding Behaviour