Mouse Alkaline Sphingomyelinase (Alk-Smase) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to Alk-Smase. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to Alk-Smase. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After the TMB substrate solution is added, only those wells that contain Alk-Smase, biotin-conjugated antibody, and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution, and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of Alk-Smase in the samples is then determined by comparing the O.D. of the samples to the standard curve.

For Use with serum, plasma, and cell culture supernatants. For Research Use Only. Not for use in diagnostic procedures.

Target Information

Choline-specific phosphodiesterase that hydrolyzes sphingomyelin releasing the ceramide and phosphocholine and therefore is involved in sphingomyelin digestion, ceramide formation, and fatty acid (FA) absorption in the gastrointestinal tract (PubMed:21177474). Has also phospholipase C activity and can also cleave phosphocholine from palmitoyl lyso-phosphatidylcholine and platelet-activating factor (PAF) leading to its inactivation. Does not have nucleotide pyrophosphatase activity (By similarity). May promote cholesterol absorption by affecting the levels of sphingomyelin derived from either diet or endogenous sources, in the intestinal lumen (PubMed:24650549).

GENE ID	238011
SWISS PROT	Q3TIW9
SYNONYMS	ENPP7; E-NPP 7; NPP-7; Ectonucleotide Pyrophosphatase/Phosphodiesterase 7; Sphingomyelin Phosphodiesterase; Intestinal alkaline sphingomyelinase

Materials Supplied

Kit Components	96 Wells Quantity/Size
Pre-coated, ready-to-use 96-well strip plate	1 plate
Plate sealer for 96 wells	2
Standard	2 tubes
Diluent buffer	1 bottle
Detection Reagent A	1 bottle
Detection Reagent B	1 bottle
TMB Substrate	1 tube
Stop Solution	1 tube
Wash Buffer (30 ℅ concentrate)	1 tube
Product data sheet	1 copy

Storage

The TMB Substrate, Wash Buffer (30X concentrate), and the Stop Solution should be stored at 4°C upon receipt, while the other items should be stored at -20°C.

Performance Characteristics

REPEATABILITY	Intra-assay Precision (Precision within an assay): 3 samples with low, middle, and high-level Alk-Smase were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level Alk-Smase were tested on 3 different plates, with 8 replicates in each plate. CV(%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%
SENSITIVITY	The minimum detectable dose was 0.051ng/mL.
ASSAY RANGE	0.156-10ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of Alk-Smase. No significant cross-reactivity or interference between Alk-Smase and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between Alk-Smase and all analogs, therefore, cross reactivity may still exist.