ELISA Kit for Ghrelin (GHRL)

MTLRP; Growth Hormone-Releasing Peptide

Specificity

This assay has high sensitivity and excellent specificity for detection of Ghrelin (GHRL).
No significant cross-reactivity or interference between Ghrelin (GHRL) and analogues was observed.

Recovery

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

Matrix Recovery range (%) Average(%)
serum(n=5) 98-105 101
EDTA plasma(n=5) 80-92 88
heparin plasma(n=5) 96-103 101

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Ghrelin (GHRL) were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level Ghrelin (GHRL) 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 Ghrelin (GHRL) 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) 93-102% 97-104% 78-89% 80-96%
EDTA plasma(n=5) 78-95% 78-92% 82-99% 80-97%
heparin plasma(n=5) 95-103% 89-96% 95-103% 84-98%

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 1×120µL Assay Diluent A 1×12mL
Detection Reagent B 1×120µL Assay Diluent B 1×12mL
TMB Substrate 1×9mL Stop Solution 1×6mL
Wash Buffer (30 × concentrate) 1×20mL Instruction manual 1

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.

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Magazine Citations
Archives of Oral Biology Expression of ghrelin in human salivary glands and its levels in saliva and serum in Chinese obese children and adolescents PubMed: 21075359
Department of Medical Biochemistry and Clinical Biochemistry Presence of adropin, nesfatin-1, apelin-12, ghrelins and salusins peptides in the milk, cheese whey and plasma of dairy cows ScienceDirect: S0196978113000533
Journal of Physiology and Pharmacology Postnatal administration of 2-oxoglutaric acid improves articular and growth plate cartilages and bone tissue morphology in pigs prenatally treated with dexamethasone PubMed: 23211309
Gene The modifying effects of fish oil on fasting ghrelin mRNA expression in weaned rats PubMed: 22842192
Hormone Research in Paediatrics Changes in Ghrelin and Obestatin Levels before and after a Meal in Children with Simple Obesity and Anorexia Pubmed: 23774845
Surgical Innovation Gastric supply manipulation to modulate ghrelin production and enhance vascularization to the cardia: proof of the concept in a porcine model Pubmed:25294792
Obesity Surgery Embolization of Arterial Gastric Supply in Obesity (EMBARGO): an Endovascular Approach in the Management of Morbid Obesity. Proof of the Concept in the Porcine Model Pubmed:25511751
Obes Surg Effect of Sleeve Gastrectomy Plus Side-to-Side Jejunoileal Anastomosis for Type 2 Diabetes Control in an Obese Rat Model PubMed: 26202420
Surg Innov Gastric Supply Manipulation to Modulate Ghrelin Production and Enhance Vascularization to the Cardia Proof of the Concept in a Porcine Model PubMed: 25294792
Nutrients.  Elevation of Fasting Ghrelin in Healthy Human Subjects Consuming a High-Salt Diet: A Novel Mechanism of Obesity? Pubmed:27240398
World Journal of Gastroenterology Effects of sleeve gastrectomy with jejuno-jejunal or jejuno-ileal loop onglycolipid metabolism in diabetic rats. pubmed:27621579
Ginekologia Polska Increased serum ghrelin in preeclampsia: is ghrelin a friend or a foe? pubmed:27321099
Gynecological Endocrinology The association between serum ghrelin levels and large-for-gestational-age fetuses in patients with gestational diabetes mellitus pubmed:27791436
Acta Diabetologica Effect of lifestyle improvement program on the biomarkersof adiposity, inflammation and gut hormones in overweight/obese Asian Indians with prediabetes pubmed:28620678
Scandinavian Journal of Clinical and Laboratory Investigation Role of serotonin hormone in weight regain after sleeve gastrectomy pubmed:29228802
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