Mouse Choline acetyltransterase (ChAT) ELISA Kit-ELISA Kit-Products-Abebio

Cat. No.	Size	Price
AE61452MO	48T	360
AE61452MO	96T	680

Technical Data

Species Reactivity

Mouse (Mus musculus)

UniProt

Q03059

Abbreviation

ChAT

Alternative Names

CMS1A; CMS1A2; acetyl CoA:choline O-acetyltransferase

Range

15.6-1000 pg/mL

Sensitivity

10.4 pg/mL

Sample Type

Serum, Plasma, Other biological fluids

Detection Method

Sandwich

Analysis Method

Quantitive

Assay Duration

1-4.5h

Sample Volume

1-200 μL

Detection Wavelengt

450 nm Test principle:

This assay employs a two-site sandwich ELISA to quantitate ChAT in Mouse serum, plasma. An antibody specific for ChAT has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any ChAT present is bound by the immobilized antibody. After removing any unbound substances, a biotin-conjugated antibody specific for ChAT is added to the wells. After washing, Streptavidin conjugated Horseradish Peroxidase (HRP) is added to the wells. Following a wash to remove any unbound avidin-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of ChAT bound in the initial step. The color development is stopped and the intensity of the color is measured.

Product Overview:

Choline acetyltransferase (EC 2.3.1.6; abbreviated ChAT) is an enzyme that is synthesized within the body of a neuron. It is then transferred to the nerve terminal via axoplasmic flow. The role of choline acetyltransferase is to join Acetyl-CoA to choline, resulting in the formation of the neurotransmitter acetylcholine. Cholinergic systems are implicated in numerous neurologic functions. Alteration in some cholinergic neurons may account for the disturbances of Alzheimer disease. The protein encoded by this gene synthesizes the neurotransmitter acetylcholine. Alternative splice variants have been found that contain alternative 5' untranslated exons. Three of the four described splice variants encode identical 69 kDa proteins while one variant encodes both the 69 kDa and a larger 82 kDa protein.

Components:

Reagents	Quantity	Reagents	Quantity
Assay plate (96 Wells)	1	Instruction manual	1
Standard (lyophilized)	2	Sample Diluent	1 x 20 mL
Biotin-Conjugate (concentrate100 x)	1 x 120 μL	Biotin-Conjugate Diluent	1 x 12 mL
Streptavidin-HRP (concentrate100 x)	1 x 120 μL	Streptavidin-HRP Diluent	1 x 12 mL
Wash Buffer (concentrate25x)	1 x 20 mL	Substrate Solution	1 x 10 mL
Stop Solution	1 x 6 mL	Adhesive Films	4

Reagents	Quantity	Reagents	Quantity
Assay plate (96 Wells)	1	Instruction manual	1
Standard (lyophilized)	2	Sample Diluent	1 x 20 mL
Biotin-Conjugate (concentrate 100 x)	1 x 120 μL	Biotin-Conjugate Diluent	1 x 20 mL
Streptavidin-HRP (concentrate 100 x)	1 x 120 μL	Streptavidin-HRP Diluent	1 x 20 mL
Wash Buffer (concentrate 25 x)	1 x 20 mL	Substrate Solution	1 x 12 mL
Stop Solution	1 x 10 mL	Adhesive Films	4

Specificity: This assay has high sensitivity and excellent specificity for detection of Mouse ChAT. No significant cross-reactivity or interference between Mouse ChAT and analogues was observed.
Recovery:

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

Sample Type	Number	Recovery range (%)	Average(%)
Serum	10	90-101	96
EDTA plasma	10	89-97	93
Heparinplasma	10	91-99	95

Sample Type	Number	Recovery range (%)	Average(%)
Serum	10	90-101	96
EDTA plasma	10	89-97	93
Heparin plasma	10	91-99	95

Precision: Intra-assay Precision (Precision within an assay) Three samples of known concentration were tested twenty times on one plate to assess intra-assay precision. Inter-assay Precision (Precision between assays) Three samples of known concentration were tested in forty separate assays to assess inter-assay precision. CV (%) = SD/meanX100 Intra-Assay: CV<8% Inter-Assay: CV<12%
Linearity: The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Mouse ChAT and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.

Sample Type	1：2	1：4	1：8	1：16
Serum	78-89%	81-99%	92-103%	95-105%
EDTA plasma	91-101%	90-98%	93-101%	91-98%
Heparin plasma	92-103%	93-102%	92-99%	91-101%

Stability: The stability of ELISA 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. The loss rate was determined by accelerated thermal degradation test. Keep the kit at 37°C for 4 and 7 days, and compare O.D.values of the kit kept at 37°C with that of at recommended temperature. (referring from China Biological Products Standard, which was calculated by the Arrhenius equation. For ELISA kit, 4 days storage at 37°C can be considered as 6 months at 2 - 8°C, which means 7 days at 37°C equaling 12 months at 2 - 8°C).
Sample collection and storage: Serum: Use a serum separator tube (SST) and allow samples to clot for two hours at room temperature or overnight at 2 - 8°C before centrifugation for 15 minutes at 1000 × g. Remove serum and assay immediately or aliquot and store samples at ≤ -20°C. Avoid repeated freeze-thaw cycles. Plasma: Collect plasma using EDTA, or heparin as an anticoagulant. Centrifuge for 15 minutes at 1000 × g at 2 - 8°C within 30 minutes of collection. Assay immediately or aliquot and store samples at ≤ -20°C. Avoid repeated freeze-thaw cycles. Other biological fluids: Centrifuge samples for 20 minutes at 1000 × g. Remove particulates and assay immediately or store samples in aliquot at -20°C or -80°C. Avoid repeated freeze/thaw cycles.
Kits storage instructions: Store at 2-8°C. Please refer to Instruction Manual.

Summary

Research topic

Bone and cartilage metabolism

Summary

Vitamin D is mainly synthesized in the skin from 7-dehydrocholesterol and is partially from dietary and supplementation origin. In the liver, Vitamin D is hydroxylated on carbon 25 to produce the intermediate 25OH Vitamin D. 25OH Vitamin D is further metabolized before it can carry out the functions of Vitamin D on intestine, kidneys, bone and other organs and tissues. This subsequent reaction takes place in the kidneys and in other tissues. Thus 25OH Vitamin D is further hydroxylated in the 1α-position to produce 1α,25-dihydroxyvitamin D (1,25(OH)2 Vitamin D). In addition to the above-mentioned tissues, placenta of pregnant women and macrophage cells in case of sarcoidis can also produce some amount of 1,25(OH)2 Vitamin D. 1,25(OH)2 Vitamin D is the active form of Vitamin D with regard to the known functions whereas 25OH Vitamin D and Vitamin D itself can be excluded as being physiologically functional. 1,25(OH)2 Vitamin D stimulates the intestinal absorption of both calcium and phosphorus. It also stimulates bone resorption and mineralization thereby preventing the development of rickets and osteomalacia. 1,25(OH)2 Vitamin D is also be active in other tissues responsible for Calcium transport (placenta, kidney, mammary gland,…) and endocrine glands such as parathyroid glands. 1,25(OH)2 Vitamin D is rapidly metabolized and its halflife is approximately 12h in plasma. Its main metabolite is calcitroic acid, a C-23 carboxylic derivative, essentially without any biological activity. In addition to this pathway, 1,25(OH)2 Vitamin D undergoes 24-hydroxylation to produce 1,24,25-trihydroxyvitamin D. This compound has less biological activity than its parent and this metabolic route is considered as a minor pathway. The levels of 1,25(OH)2 Vitamin D in plasma or serum is 100 to 1000 less than that of 25OH Vitamin D. Due to its low concentrations and the presence of many similar metabolites, the measurement of 1,25(OH)2 Vitamin D requires extraction and separation by chromatography.

Product References (1)

References

Parfieniuk-Kowerda A, Świderska M, Rogalska M, Maciaszek M, Jaroszewicz J,Flisiak R. Chronic hepatitis B virus infection is associated with decreased serum25(OH)D concentration in non-cirrhotic patients. Clin Exp Hepatol. 2019Mar;5(1):75-80. doi: 10.5114/ceh.2019.83160. Epub 2019 Feb 20. PubMed PMID:30915410; PubMed Central PMCID: PMC6431090. See more on PubMed

Summary References (1)

References to 1,25OH2 Vitamin D

Bouillon RA, Auwerx JH, Lissens WD, Pelemans WK. Vitamin D status in the elderly: seasonal substrate deficiency causes 1,25-dihydroxycholecalciferol deficiency. Am J Clin Nutr. 1987 Apr;45 (4):755-63

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Mouse Choline acetyltransterase (ChAT) ELISA Kit

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References to 1,25OH2 Vitamin D

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