Melatonin ELISA Kit

Catalog No. ABIN6574097

Product Details Melatonin ELISA Kit

Target: Melatonin (MT)

Reactivity: Various Species

Detection Method: Colorimetric

Method Type: Competition ELISA

Detection Range: 12.35 pg/mL - 1000 pg/mL

Minimum Detection Limit: 12.35 pg/mL

Application: ELISA

Purpose: The kit is a competitive inhibition enzyme immunoassay technique for the in vitro quantitative measurement of melatonin in serum, plasma.

We offer validation data (WB) for the kit components. So you can be sure to order a reliable ELISA kit product composed of high quality reagents.

Sample Type: Plasma, Serum

Analytical Method: Quantitative

Specificity: This assay has high sensitivity and excellent specificity for detection of Melatonin (MT)

Cross-Reactivity (Details): No significant cross-reactivity or interference between Melatonin (MT) and analogues was observed.

Sensitivity: 4.63 pg/mL

Components:

• 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

Application Details

Comment:

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.

Sample Volume: 50 μL

Assay Time: 2 h

Plate: Pre-coated

Protocol:

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.

Reagent Preparation:

1. Bring all kit components and samples to room temperature (18-25 °C) before use. If the kit will not be used up in one time, please only take out strips and reagents for present experiment, and leave the remaining strips and reagents in required condition.
2. Standard - Reconstitute the Standard with 2.0 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 2,000pg/mL. Please firstly dilute the stock solution to 1,000pg/mL and the diluted standard serves as the highest standard (1,000pg/mL). Then prepare 5 tubes containing 0.6 mL Standard Diluent and produce a triple dilution series according to the picture shown below. 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 - If lyophilized reconstitute the Detection Reagent A with 150μL of Reagent Diluent, kept for 10 minutes at room temperature, shake gently (not to foam). Briefly spin or centrifuge the stock Detection A and Detection B before use. Dilute them to the working concentration 100-fold with Assay Diluent A and B, respectively.
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 one 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.

Sample Preparation:

• It is recommended to use fresh samples without long storage, otherwise protein degradation and denaturationmay occur in these samples, leading to false results. Samples should therefore be stored for a short periodat 2 - 8 °C or aliquoted at -20 °C (≤1 month) or -80 °C (≤ 3 months). Repeated freeze-thawcycles should be avoided. Prior to assay, the frozen samples should be slowly thawed and centrifuged toremove precipitates.
• If the sample type is not specified in the instructions, a preliminary test is necessary to determinecompatibility with the kit.
• If a lysis buffer is used to prepare tissue homogenates or cell culture supernatant, there is a possibilityof 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 therange of the standard curve, the optimal sample dilution for the particular experiment has to be determined.Samples should then be diluted with PBS (pH =7.0-7.2).

Assay Precision: 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%

Restrictions: For Research Use only

Handling

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

Storage: 4 °C/-20 °C

Storage Comment:

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.

.

Expiry Date: 6 months

References for Melatonin ELISA Kit (ABIN6574097)

Szewczyk-Golec, Rajewski, Gackowski, Mila-Kierzenkowska, Wesołowski, Sutkowy, Pawłowska, Woźniak: "Melatonin Supplementation Lowers Oxidative Stress and Regulates Adipokines in Obese Patients on a Calorie-Restricted Diet." in: Oxidative medicine and cellular longevity, Vol. 2017, pp. 8494107, (2018) (PubMed).

Zhang, Chen, Cao, Dong, Wang, Chen: "Melatonin modulates monochromatic light-induced melatonin receptor expression in the hypothalamus of chicks." in: Acta histochemica, Vol. 119, Issue 7, pp. 733-739, (2018) (PubMed).

Zhang, Chen, Cao, Dong, Wang, Hu, Chen: "Green light inhibits GnRH-I expression by stimulating the melatonin-GnIH pathway in the chick brain." in: Journal of neuroendocrinology, Vol. 29, Issue 5, (2017) (PubMed).

Özdem, Kırzıoğlu, Yılmaz, Vural, Fentoğlu, Uz, Koçak, Yiğit: "Antioxidant effects of melatonin in heart tissue after induction of experimental periodontitis in rats." in: Journal of oral science, Vol. 59, Issue 1, pp. 23-29, (2017) (PubMed).

Chang, Lin, Zhuang, Huang, Lin, Jea, Fan, Lee: "Effect of Monochromic Light-emitting Diode Light with Different Color on the Growth and Reproductive Performances of Breeder Geese." in: Asian-Australasian journal of animal sciences, Vol. 29, Issue 6, pp. 830-7, (2016) (PubMed).

Chen, Reheman, Cao, Wang, Dong, Zhang, Chen: "Effect of melatonin on monochromatic light-induced T-lymphocyte proliferation in the thymus of chickens." in: Journal of photochemistry and photobiology. B, Biology, Vol. 161, pp. 9-16, (2016) (PubMed).

Zhang, Cao, Wang, Dong, Chen: "Melatonin modulates monochromatic light-induced GHRH expression in the hypothalamus and GH secretion in chicks." in: Acta histochemica, Vol. 118, Issue 3, pp. 286-92, (2016) (PubMed).

Roy, Singh, Kumar, Kumar: "Effect of extended photoperiod during winter on growth and onset of puberty in Murrah buffalo heifers." in: Veterinary world, Vol. 9, Issue 2, pp. 216-21, (2016) (PubMed).

Jiang, Wang, Cao, Dong, Chen: "Role of monochromatic light on daily variation of clock gene expression in the pineal gland of chick." in: Journal of photochemistry and photobiology. B, Biology, Vol. 164, pp. 57-64, (2016) (PubMed).

Lee, OHare, Chen, Chang, Yang, Liu, Lin: "Molecularly imprinted electrochemical sensing of urinary melatonin in a microfluidic system." in: Biomicrofluidics, Vol. 8, Issue 5, pp. 054115, (2015) (PubMed).

Guo, Dong, Cao, Wang, Zhang, Chen: "Developmental changes of melatonin receptor expression in the spleen of the chicken, Gallus domesticus." in: Acta histochemica, Vol. 117, Issue 6, pp. 559-65, (2015) (PubMed).

Singh, Ghosh, Basu, Haldar: "Daily variation in melatonin level, antioxidant activity and general immune response of peripheral blood mononuclear cells and lymphoid tissues of Indian goat Capra hircus during summer and winter." in: Indian journal of experimental biology, Vol. 52, Issue 5, pp. 467-77, (2014) (PubMed).

Jain, Jain, Haldar, Singh, Jain: "Melatonin and its correlation with testosterone in polycystic ovarian syndrome." in: Journal of human reproductive sciences, Vol. 6, Issue 4, pp. 253-8, (2014) (PubMed).

Vishwas, Haldar: "MT1 receptor expression and AA-NAT activity in lymphatic tissue following melatonin administration in male golden hamster." in: International immunopharmacology, Vol. 22, Issue 1, pp. 258-65, (2014) (PubMed).

Cernysiov, Bozaite, Mauricas, Girkontaite: "Influence of circadian time and lighting conditions on expression of melatonin receptors 1 and 2 in murine lymphocytes." in: In vivo (Athens, Greece), Vol. 28, Issue 5, pp. 831-5, (2014) (PubMed).

Mukherjee, Haldar: "Photoperiodic regulation of melatonin membrane receptor (MT1R) expression and steroidogenesis in testis of adult golden hamster, Mesocricetus auratus." in: Journal of photochemistry and photobiology. B, Biology, Vol. 140, pp. 374-80, (2014) (PubMed).

Mukherjee, Haldar, Vishwas: "Melatonin prevents dexamethasone-induced testicular oxidative stress and germ cell apoptosis in golden hamster, Mesocricetus auratus." in: Andrologia, (2014) (PubMed).

Vishwas, Mukherjee, Haldar, Dash, Nayak: "Improvement of oxidative stress and immunity by melatonin: an age dependent study in golden hamster." in: Experimental gerontology, Vol. 48, Issue 2, pp. 168-82, (2013) (PubMed).

Vishwas, Haldar: "Photoperiodic induced melatonin regulates immunity and expression pattern of melatonin receptor MT1 in spleen and bone marrow mononuclear cells of male golden hamster." in: Journal of photochemistry and photobiology. B, Biology, Vol. 128, pp. 107-14, (2013) (PubMed).

Jin, Choi, Kim, Pack, Kim, Lee: "Effects of simultaneous combined exposure to CDMA and WCDMA electromagnetic fields on serum hormone levels in rats." in: Journal of radiation research, Vol. 54, Issue 3, pp. 430-7, (2013) (PubMed).

Target Details for Melatonin

Target: Melatonin (MT)

Abstract: MT Products

Target Type: Chemical