Mouse Transforming protein RhoA (Rhoa) ELISA Kit (MOEB2411)
- SKU:
- MOEB2411
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- Q9QUI0
- ELISA Type:
- Sandwich
- Reactivity:
- Mouse
Description
Mouse Transforming protein RhoA (Rhoa) ELISA Kit
The Mouse Transforming Protein RHOA (RhoA) ELISA Kit is a powerful tool for the detection and quantification of RhoA levels in mouse serum, plasma, and cell culture supernatants. With its high sensitivity and specificity, this kit ensures accurate and reliable results, making it suitable for a wide range of research applications.RhoA is a key regulator of cell motility and shape, playing a critical role in various cellular processes such as cytoskeletal dynamics, cell adhesion, and cell contraction. Dysregulation of RhoA activity has been implicated in diseases like cancer, cardiovascular disorders, and neurological conditions, highlighting its importance as a potential therapeutic target.
By utilizing the Mouse Transforming Protein RHOA (RhoA) ELISA Kit, researchers can gain valuable insights into the role of RhoA in disease pathogenesis and explore its potential as a biomarker for disease diagnosis and treatment. With its user-friendly protocol and reliable performance, this kit is an essential tool for advancing scientific knowledge in the field of molecular biology and drug discovery.
| Product Name: | Mouse Transforming protein RhoA (Rhoa) ELISA Kit |
| SKU: | MOEB2411 |
| Size: | 96T |
| Target: | Mouse Transforming protein RhoA (Rhoa) |
| Synonyms: | Arha, Arha2 |
| Assay Type: | Sandwich |
| Detection Method: | ELISA |
| Reactivity: | Mouse |
| Detection Range: | 0.156-10ng/mL |
| Sensitivity: | 0.068ng/mL |
| Intra CV: | 4.8% | ||||||||||||||||||||
| Inter CV: | 7.7% | ||||||||||||||||||||
| Linearity: |
| ||||||||||||||||||||
| Recovery: |
| ||||||||||||||||||||
| Function: | Regulates a signal transduction pathway linking plasma membrane receptors to the assembly of focal adhesions and actin stress fibers. Involved in a microtubule-dependent signal that is required for the myosin contractile ring formation during cell cycle cytokinesis. Plays an essential role in cleavage furrow formation. May be an activator of PLCE1. Activated by ARHGEF2, which promotes the exchange of GDP for GTP. Essential for the SPATA13-mediated regulation of cell migration and adhesion assembly and disassembly. The MEMO1-RHOA-DIAPH1 signaling pathway plays an important role in ERBB2-dependent stabilization of microtubules at the cell cortex. It controls the localization of APC and CLASP2 to the cell membrane, via the regulation of GSK3B activity. In turn, membrane-bound APC allows the localization of the MACF1 to the cell membrane, which is required for microtubule capture and stabilization (By similarity). Required for the apical junction formation of keratinocyte cell-cell adhesion. Regulates KCNA2 potassium channel activity by reducing its location at the cell surface in response to CHRM1 activation; promotes KCNA2 endocytosis (PubMed:9635436). |
| Uniprot: | Q9QUI0 |
| Sample Type: | Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids |
| Specificity: | Natural and recombinant mouse Transforming protein RhoA |
| Sub Unit: | Binds PRKCL1, ROCK1 and ROCK2. Interacts with ARHGEF2 and ARHGEF3. Interacts with PLCE1 and AKAP13. Interacts with DIAPH1. Interacts (in the constitutively activated, GTP-bound form) with DGKQ. Interacts with RACK1; enhances RHOA activation. Interacts with PKP4; the interaction is detected at the midbody. Interacts (GTP-bound form preferentially) with PKN2; the interaction stimulates autophosphorylation and phosphorylation of PKN2 (By similarity). Interacts with NET1, ARHGEF28 and RTKN. Interacts with ARHGDIA; this interaction inactivates and stabilizes RHOA. Interacts with ARHGDIB (By similarity). Interacts (GTP-bound form) with KCNA2 (via cytoplasmic N-terminal domain) (PubMed:9635436). |
| Research Area: | Signal Transduction |
| Subcellular Location: | Cell membrane Lipid-anchor Cytoplasmic side Cytoplasm Cytoskeleton Cleavage furrow Cytoplasm Cell cortex Midbody Cell projection Lamellipodium Translocates to the equatorial region before furrow formation in a ECT2-dependent manner. Localizes to the equatorial cell cortex (at the site of the presumptive furrow) in early anaphase in a activated form and in a myosin- and actin-independent manner (By similarity). Localized to cell-cell contacts in calcium-treated keratinocytes. |
| Storage: | Please see kit components below for exact storage details |
| Note: | For research use only |
| UniProt Protein Function: | RHOA: a small G protein of the Rho family. Regulates a signal transduction pathway linking plasma membrane receptors to the assembly of focal adhesions and actin stress fibers. Controls the reorganization of actins into podosomes. Serves as a target for the yopT cysteine peptidase from Yersinia pestis, vector of the plague, and Yersinia pseudotuberculosis, which causes gastrointestinal disorders. |
| UniProt Protein Details: | Protein type:G protein, monomeric; G protein; Motility/polarity/chemotaxis; G protein, monomeric, Rho; Oncoprotein Cellular Component: apical junction complex; focal adhesion; mitochondrion; cell cortex; cytosol; cytoskeleton; cell projection; membrane; lamellipodium; axon; cytoplasm; plasma membrane; intracellular; nucleus Molecular Function:GTPase activity; protein domain specific binding; protein binding; GDP binding; myosin binding; GTP binding; Rho GDP-dissociation inhibitor binding; nucleotide binding Biological Process: radial glial cell differentiation in the forebrain; metabolic process; cell-matrix adhesion; cell morphogenesis; regulation of dendrite development; positive regulation of caspase activity; negative chemotaxis; regulation of cell migration; Rho protein signal transduction; regulation of osteoblast proliferation; small GTPase mediated signal transduction; positive regulation of neuron apoptosis; positive regulation of stress fiber formation; stress-activated protein kinase signaling pathway; negative regulation of neuron apoptosis; positive regulation of smooth muscle contraction; cell adhesion; cell differentiation; response to drug; integrin-mediated signaling pathway; positive regulation of cytokinesis; negative regulation of steroid hormone receptor signaling pathway; skeletal muscle development; apical junction assembly; cytoskeleton organization and biogenesis; regulation of calcium ion transport; negative regulation of I-kappaB kinase/NF-kappaB cascade; cell cycle; positive regulation of cell growth; regulation of transcription from RNA polymerase II promoter; negative regulation of neuron differentiation; cerebral cortex cell migration; positive regulation of actin filament polymerization; cell division; androgen receptor signaling pathway; stress fiber formation; positive regulation of neuron differentiation; actin cytoskeleton organization and biogenesis |
| NCBI Summary: | This gene encodes a member of the Rho family of small GTPases, which cycle between inactive GDP-bound and active GTP-bound states and function as molecular switches in signal transduction cascades. Rho proteins promote reorganization of the actin cytoskeleton and regulate cell shape, attachment, and motility. Overexpression of this gene is associated with tumor cell proliferation and metastasis. Multiple alternatively spliced variants have been identified. [provided by RefSeq, Sep 2015] |
| UniProt Code: | Q9QUI0 |
| NCBI GenInfo Identifier: | 31542143 |
| NCBI Gene ID: | 11848 |
| NCBI Accession: | NP_058082.2 |
| UniProt Secondary Accession: | Q9QUI0,O88336, |
| UniProt Related Accession: | Q9QUI0 |
| Molecular Weight: | 21,782 Da |
| NCBI Full Name: | transforming protein RhoA |
| NCBI Synonym Full Names: | ras homolog gene family, member A |
| NCBI Official Symbol: | Rhoa |
| NCBI Official Synonym Symbols: | Arha; Arha1; Arha2 |
| NCBI Protein Information: | transforming protein RhoA |
| UniProt Protein Name: | Transforming protein RhoA |
| Protein Family: | GTP-binding protein |
| UniProt Gene Name: | Rhoa |
| UniProt Entry Name: | RHOA_MOUSE |
| 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. |
Related Products
Human Transforming protein RhoA (RHOA) ELISA Kit (HUEB2667)
Dog Transforming protein RhoA (RHOA) ELISA Kit (CNEB0371)
Bovine Transforming protein RhoA (RHOA) ELISA Kit (BOEB1154)
Human RhoA ELISA Kit
Human Ras Homolog Gene Family, Member A (RHOA) ELISA Kit
