Description
Rat Aldehyde dehydrogenase, mitochondrial (Aldh2) ELISA Kit
The Rat Aldehyde Dehydrogenase Mitochondrial (ALDH2) ELISA Kit is specifically designed for the accurate quantification of ALDH2 levels in rat samples including serum, plasma, and tissue lysates. This kit offers high sensitivity and specificity, ensuring precise and reproducible results for a variety of research applications.ALDH2 is a key enzyme responsible for the oxidation of aldehydes, playing a critical role in detoxifying harmful compounds and maintaining cellular homeostasis.
Dysregulation of ALDH2 has been linked to various diseases including alcohol-induced liver damage, cardiovascular diseases, and cancer, making it a valuable biomarker for studying these conditions and exploring potential therapeutic interventions.Overall, the Rat Aldehyde Dehydrogenase Mitochondrial (ALDH2) ELISA Kit provides researchers with a reliable tool for investigating the role of ALDH2 in disease pathogenesis and identifying new avenues for treatment strategies.
| Product Name: | Rat Aldehyde dehydrogenase, mitochondrial (Aldh2) ELISA Kit |
| SKU: | RTEB1328 |
| Size: | 96T |
| Target: | Rat Aldehyde dehydrogenase, mitochondrial (Aldh2) |
| Synonyms: | ALDH class 2, ALDH-E2, ALDH1 |
| Assay Type: | Sandwich |
| Detection Method: | ELISA |
| Reactivity: | Rat |
| Detection Range: | 0.312-20ng/mL |
| Sensitivity: | 0.186ng/mL |
| Intra CV: | 4.7% | ||||||||||||||||||||
| Inter CV: | 7.1% | ||||||||||||||||||||
| Linearity: |
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| Recovery: |
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| Uniprot: | P11884 |
| Sample Type: | Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids |
| Specificity: | Natural and recombinant rat Aldehyde dehydrogenase, mitochondrial |
| Sub Unit: | Homotetramer. |
| Research Area: | Signal Transduction |
| Subcellular Location: | Mitochondrion matrix |
| Storage: | Please see kit components below for exact storage details |
| Note: | For research use only |
| UniProt Protein Function: | ALDH2: This protein belongs to the aldehyde dehydrogenase family of proteins. Aldehyde dehydrogenase is the second enzyme of the major oxidative pathway of alcohol metabolism. Two major liver isoforms of aldehyde dehydrogenase, cytosolic and mitochondrial, can be distinguished by their electrophoretic mobilities, kinetic properties, and subcellular localizations. Most Caucasians have two major isozymes, while approximately 50% of Orientals have the cytosolic isozyme but not the mitochondrial isozyme. A remarkably higher frequency of acute alcohol intoxication among Orientals than among Caucasians could be related to the absence of a catalytically active form of the mitochondrial isozyme. The increased exposure to acetaldehyde in individuals with the catalytically inactive form may also confer greater susceptibility to many types of cancer. This gene encodes a mitochondrial isoform, which has a low Km for acetaldehydes, and is localized in mitochondrial matrix. Alternative splicing results in multiple transcript variants encoding distinct isoforms.[provided by RefSeq, Mar 2011] |
| UniProt Protein Details: | Protein type:Amino Acid Metabolism - arginine and proline; Secondary Metabolites Metabolism - limonene and pinene degradation; Carbohydrate Metabolism - ascorbate and aldarate; Carbohydrate Metabolism - glycolysis and gluconeogenesis; Carbohydrate Metabolism - propanoate; Oxidoreductase; EC 1.2.1.3; Other Amino Acids Metabolism - beta-alanine; Carbohydrate Metabolism - pyruvate; Mitochondrial; Amino Acid Metabolism - lysine degradation; Amino Acid Metabolism - tryptophan; Amino Acid Metabolism - valine, leucine and isoleucine degradation; Amino Acid Metabolism - histidine; Lipid Metabolism - glycerolipid; Lipid Metabolism - fatty acid; Carbohydrate Metabolism - butanoate Cellular Component: mitochondrial matrix; mitochondrion Molecular Function:aldehyde dehydrogenase (NAD) activity; identical protein binding; oxidoreductase activity, acting on the aldehyde or oxo group of donors, NAD or NADP as acceptor; protein binding Biological Process: cellular response to hormone stimulus; ethanol catabolic process; liver development; negative regulation of apoptosis; response to estradiol stimulus; response to hyperoxia; response to lipopolysaccharide; response to nicotine; response to progesterone stimulus; response to testosterone stimulus |
| UniProt Code: | P11884 |
| NCBI GenInfo Identifier: | 14192933 |
| NCBI Gene ID: | 29539 |
| NCBI Accession: | NP_115792.1 |
| UniProt Secondary Accession: | P11884,Q6Q288, Q6Q289, Q6Q290, Q8K3V8, Q91ZD7, |
| UniProt Related Accession: | P11884 |
| Molecular Weight: | 56,488 Da |
| NCBI Full Name: | aldehyde dehydrogenase, mitochondrial |
| NCBI Synonym Full Names: | aldehyde dehydrogenase 2 family (mitochondrial) |
| NCBI Official Symbol: | Aldh2 |
| NCBI Protein Information: | aldehyde dehydrogenase, mitochondrial |
| UniProt Protein Name: | Aldehyde dehydrogenase, mitochondrial |
| UniProt Synonym Protein Names: | ALDH class 2; ALDH-E2; ALDH1 |
| Protein Family: | NADP-dependent fatty aldehyde dehydrogenase |
| UniProt Gene Name: | Aldh2 |
| UniProt Entry Name: | ALDH2_RAT |
| Component | Quantity (96 Assays) | Storage |
| ELISA Microplate (Dismountable) | 8×12 strips | -20°C |
| Lyophilized Standard | 2 | -20°C |
| Sample Diluent | 20ml | -20°C |
| Assay Diluent A | 10mL | -20°C |
| Assay Diluent B | 10mL | -20°C |
| Detection Reagent A | 120µL | -20°C |
| Detection Reagent B | 120µL | -20°C |
| Wash Buffer | 30mL | 4°C |
| Substrate | 10mL | 4°C |
| Stop Solution | 10mL | 4°C |
| Plate Sealer | 5 | - |
Other materials and equipment required:
- Microplate reader with 450 nm wavelength filter
- Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
- Incubator
- Deionized or distilled water
- Absorbent paper
- Buffer resevoir
*Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
Allow all reagents to reach room temperature (Please do not dissolve the reagents at 37°C directly). All the reagents should be mixed thoroughly by gently swirling before pipetting. Avoid foaming. Keep appropriate numbers of strips for 1 experiment and remove extra strips from microtiter plate. Removed strips should be resealed and stored at -20°C until the kits expiry date. Prepare all reagents, working standards and samples as directed in the previous sections. Please predict the concentration before assaying. If values for these are not within the range of the standard curve, users must determine the optimal sample dilutions for their experiments. We recommend running all samples in duplicate.
| Step | |
| 1. | Add Sample: Add 100µL of Standard, Blank, or Sample per well. The blank well is added with Sample diluent. Solutions are added to the bottom of micro ELISA plate well, avoid inside wall touching and foaming as possible. Mix it gently. Cover the plate with sealer we provided. Incubate for 120 minutes at 37°C. |
| 2. | Remove the liquid from each well, don't wash. Add 100µL of Detection Reagent A working solution to each well. Cover with the Plate sealer. Gently tap the plate to ensure thorough mixing. Incubate for 1 hour at 37°C. Note: if Detection Reagent A appears cloudy warm to room temperature until solution is uniform. |
| 3. | Aspirate each well and wash, repeating the process three times. Wash by filling each well with Wash Buffer (approximately 400µL) (a squirt bottle, multi-channel pipette,manifold dispenser or automated washer are needed). Complete removal of liquid at each step is essential. After the last wash, completely remove remaining Wash Buffer by aspirating or decanting. Invert the plate and pat it against thick clean absorbent paper. |
| 4. | Add 100µL of Detection Reagent B working solution to each well. Cover with the Plate sealer. Incubate for 60 minutes at 37°C. |
| 5. | Repeat the wash process for five times as conducted in step 3. |
| 6. | Add 90µL of Substrate Solution to each well. Cover with a new Plate sealer and incubate for 10-20 minutes at 37°C. Protect the plate from light. The reaction time can be shortened or extended according to the actual color change, but this should not exceed more than 30 minutes. When apparent gradient appears in standard wells, user should terminatethe reaction. |
| 7. | Add 50µL of Stop Solution to each well. If color change does not appear uniform, gently tap the plate to ensure thorough mixing. |
| 8. | Determine the optical density (OD value) of each well at once, using a micro-plate reader set to 450 nm. User should open the micro-plate reader in advance, preheat the instrument, and set the testing parameters. |
| 9. | After experiment, store all reagents according to the specified storage temperature respectively until their expiry. |
When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.
| Sample Type | Protocol |
| Serum | If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. |
| Plasma | Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit. |
| Urine & Cerebrospinal Fluid | Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid. |
| Cell culture supernatant | Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately. |
| Cell lysates | Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C. |
| Tissue homogenates | The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C. |
| Tissue lysates | Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C. |
| Breast Milk | Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles. |
