Rat Signaling ELISA Kits 4
Rat ACE (Angiotensin I Converting Enzyme) ELISA Kit (RTES01029)
- SKU:
- RTES01029
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- P47820
- Sensitivity:
- 0.19ng/mL
- Range:
- 0.31-20ng/mL
- ELISA Type:
- Sandwich
- Synonyms:
- ACE1, CD143, DCP, DCP1, ICH, MVCD3
- Reactivity:
- Rat
- Sample Type:
- Serum, plasma and other biological fluids
Description
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Rat |
| Detection Method: | Colormetric |
| Detection Range: | 0.31-20 ng/mL |
| Sensitivity: | 0.19 ng/mL |
| Sample Volume Required Per Well: | 100µL |
| Sample Type: | Serum, plasma and other biological fluids |
| Specificity: | This kit recognizes Rat ACE in samples. No significant cross-reactivity or interference between Rat ACE and analogues was observed. |
This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Rat ACE. Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Rat ACE and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Rat ACE, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Rat ACE. The concentration of Rat ACE in samples can be calculated by comparing the OD of the samples to the standard curve.
| UniProt Protein Function: | ACE: Converts angiotensin I to angiotensin II by release of the terminal His-Leu, this results in an increase of the vasoconstrictor activity of angiotensin. Also able to inactivate bradykinin, a potent vasodilator. Has also a glycosidase activity which releases GPI-anchored proteins from the membrane by cleaving the mannose linkage in the GPI moiety. Genetic variations in ACE may be a cause of susceptibility to ischemic stroke (ISCHSTR); also known as cerebrovascular accident or cerebral infarction. A stroke is an acute neurologic event leading to death of neural tissue of the brain and resulting in loss of motor, sensory and/or cognitive function. Ischemic strokes, resulting from vascular occlusion, is considered to be a highly complex disease consisting of a group of heterogeneous disorders with multiple genetic and environmental risk factors. Defects in ACE are a cause of renal tubular dysgenesis (RTD). RTD is an autosomal recessive severe disorder of renal tubular development characterized by persistent fetal anuria and perinatal death, probably due to pulmonary hypoplasia from early-onset oligohydramnios (the Potter phenotype). Genetic variations in ACE are associated with susceptibility to microvascular complications of diabetes type 3 (MVCD3). These are pathological conditions that develop in numerous tissues and organs as a consequence of diabetes mellitus. They include diabetic retinopathy, diabetic nephropathy leading to end-stage renal disease, and diabetic neuropathy. Diabetic retinopathy remains the major cause of new- onset blindness among diabetic adults. It is characterized by vascular permeability and increased tissue ischemia and angiogenesis. Defects in ACE are a cause of susceptibility to intracerebral hemorrhage (ICH). A pathological condition characterized by bleeding into one or both cerebral hemispheres including the basal ganglia and the cerebral cortex. It is often associated with hypertension and craniocerebral trauma. Intracerebral bleeding is a common cause of stroke. Belongs to the peptidase M2 family. 4 isoforms of the human protein are produced by alternative splicing. |
| UniProt Protein Details: | Protein type:Membrane protein, integral; EC 3. 4. 15. 1; Protease Cellular Component: basal plasma membrane; brush border membrane; cytoplasm; endosome; external side of plasma membrane; extracellular space; integral to membrane; lysosome; plasma membrane; vesicle Molecular Function:actin binding; bradykinin receptor binding; carboxypeptidase activity; chloride ion binding; drug binding; endopeptidase activity; exopeptidase activity; metal ion binding; metallopeptidase activity; mitogen-activated protein kinase binding; mitogen-activated protein kinase kinase binding; peptidase activity; peptidyl-dipeptidase activity; tripeptidyl-peptidase activity; zinc ion binding Biological Process: aging; alveolus development; arachidonic acid secretion; beta-amyloid metabolic process; brain development; eating behavior; female pregnancy; heart contraction; hormone catabolic process; kidney development; lung development; male gonad development; negative regulation of protein binding; neutrophil mediated immunity; organ regeneration; peptide catabolic process; peptide metabolic process; positive regulation of apoptosis; positive regulation of inflammatory response; positive regulation of neurogenesis; positive regulation of protein binding; positive regulation of systemic arterial blood pressure; proteolysis; regulation of angiotensin metabolic process; regulation of blood pressure; regulation of smooth muscle cell migration; regulation of systemic arterial blood pressure by renin-angiotensin; response to drug; response to hypoxia; response to lipopolysaccharide; response to nutrient levels; sensory perception of pain; spermatogenesis; vasoconstriction |
| NCBI Summary: | catalyzes the conversion of angiotensin I to angiotensin II; plays a role in regulation of blood pressure [RGD, Feb 2006] |
| UniProt Code: | P47820 |
| NCBI GenInfo Identifier: | 6978757 |
| NCBI Gene ID: | 24310 |
| NCBI Accession: | NP_036676. 1 |
| UniProt Related Accession: | P47820 |
| Molecular Weight: | |
| NCBI Full Name: | angiotensin-converting enzyme |
| NCBI Synonym Full Names: | angiotensin I converting enzyme |
| NCBI Official Symbol: | Ace |
| NCBI Official Synonym Symbols: | Dcp1; CD143; StsRR92 |
| NCBI Protein Information: | angiotensin-converting enzyme |
| UniProt Protein Name: | Angiotensin-converting enzyme |
| UniProt Synonym Protein Names: | Dipeptidyl carboxypeptidase I; Kininase II; CD_antigen: CD143 |
| Protein Family: | Acetylcholinesterase |
| UniProt Gene Name: | Ace |
| UniProt Entry Name: | ACE_RAT |
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e. g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
| Concentration (ng/mL) | O.D | Average | Corrected |
| 20 | 2.333 2.377 | 2.355 | 2.275 |
| 10 | 1.686 1.69 | 1.688 | 1.608 |
| 5 | 0.983 0.981 | 0.982 | 0.902 |
| 2.5 | 0.418 0.45 | 0.434 | 0.354 |
| 1.25 | 0.259 0.255 | 0.257 | 0.177 |
| 0.63 | 0.192 0.17 | 0.181 | 0.101 |
| 0.31 | 0.128 0.134 | 0.131 | 0.051 |
| 0 | 0.077 0.083 | 0.08 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Rat ACE were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Rat ACE were tested on 3 different plates, 20 replicates in each plate.
| Intra-assay Precision | Inter-assay Precision | |||||
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean (ng/mL) | 1.10 | 3.06 | 9.84 | 1.20 | 3.18 | 9.23 |
| Standard deviation | 0.06 | 0.13 | 0.31 | 0.07 | 0.18 | 0.30 |
| C V (%) | 5.45 | 4.25 | 3.15 | 5.83 | 5.66 | 3.25 |
Recovery
The recovery of Rat ACE spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
| Sample Type | Range (%) | Average Recovery (%) |
| Serum (n=5) | 93-107 | 101 |
| EDTA plasma (n=5) | 95-111 | 102 |
| Cell culture media (n=5) | 87-103 | 94 |
Linearity
Samples were spiked with high concentrations of Rat ACE and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
| Serum (n=5) | EDTA plasma (n=5) | Cell culture media (n=5) | ||
| 1:2 | Range (%) | 93-109 | 95-110 | 93-110 |
| Average (%) | 100 | 100 | 100 | |
| 1:4 | Range (%) | 93-108 | 82-93 | 84-96 |
| Average (%) | 99 | 88 | 89 | |
| 1:8 | Range (%) | 95-108 | 83-94 | 82-94 |
| Average (%) | 100 | 88 | 88 | |
| 1:16 | Range (%) | 88-100 | 80-91 | 83-97 |
| Average (%) | 94 | 86 | 89 |
An unopened kit can be stored at 4°C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
| Item | Specifications | Storage |
| Micro ELISA Plate(Dismountable) | 8 wells ×12 strips | -20°C, 6 months |
| Reference Standard | 2 vials | |
| Concentrated Biotinylated Detection Ab (100×) | 1 vial, 120 µL | |
| Concentrated HRP Conjugate (100×) | 1 vial, 120 µL | -20°C(shading light), 6 months |
| Reference Standard & Sample Diluent | 1 vial, 20 mL | 4°C, 6 months |
| Biotinylated Detection Ab Diluent | 1 vial, 14 mL | |
| HRP Conjugate Diluent | 1 vial, 14 mL | |
| Concentrated Wash Buffer (25×) | 1 vial, 30 mL | |
| Substrate Reagent | 1 vial, 10 mL | 4°C(shading light) |
| Stop Solution | 1 vial, 10 mL | 4°C |
| Plate Sealer | 5 pieces | |
| Product Description | 1 copy | |
| Certificate of Analysis | 1 copy |
- Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- Aliquot 100µl of standard solutions into the standard wells.
- Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix. Incubate for 1 hour at 37°C.
- Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- Add 90µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37°C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
- Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.
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