Description
Human Mothers against decapentaplegic homolog 1 (SMAD1) ELISA Kit
The Human Mothers Against Decapentaplegic Homolog 1 (SMAD1) ELISA Kit is a dependable tool for detecting levels of SMAD1 in human samples such as serum, plasma, and cell culture supernatants. With its high sensitivity and specificity, this kit provides accurate and consistent results, making it suitable for various research applications.SMAD1 is a key protein involved in multiple cellular processes, including cell growth, differentiation, and development.
Dysregulation of SMAD1 signaling has been linked to various diseases, such as cancer, fibrosis, and developmental disorders, highlighting its importance as a potential therapeutic target and biomarker.By utilizing the Human SMAD1 ELISA Kit, researchers can gain valuable insights into the role of SMAD1 in disease pathology, paving the way for new diagnostic and therapeutic strategies. Elevate your research with this advanced ELISA kit from AssayGenie.
| Product Name: | Human Mothers against decapentaplegic homolog 1 (SMAD1) ELISA Kit |
| SKU: | HUEB0779 |
| Size: | 96T |
| Target: | Human Mothers against decapentaplegic homolog 1 (SMAD1) |
| Synonyms: | JV4-1, Mad-related protein 1, SMAD family member 1, Transforming growth factor-beta-signaling protein 1, SMAD 1, BSP-1, MAD homolog 1, BSP1, MADH1, MADR1 |
| Assay Type: | Sandwich |
| Detection Method: | ELISA |
| Reactivity: | Human |
| Detection Range: | 15.6-1000pg/mL |
| Sensitivity: | 8.9pg/mL |
| Intra CV: | 5.5% | ||||||||||||||||||||
| Inter CV: | 9.8% | ||||||||||||||||||||
| Linearity: |
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| Recovery: |
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| Function: | Transcriptional modulator activated by BMP (bone morphogenetic proteins) type 1 receptor kinase. SMAD1 is a receptor-regulated SMAD (R-SMAD). SMAD1/OAZ1/PSMB4 complex mediates the degradation of the CREBBP/EP300 repressor SNIP1. May act synergistically with SMAD4 and YY1 in bone morphogenetic protein (BMP)-mediated cardiac-specific gene expression. |
| Uniprot: | Q15797 |
| Sample Type: | Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids |
| Specificity: | Natural and recombinant human Mothers against decapentaplegic homolog 1 |
| Sub Unit: | Found in a complex with SMAD4 and YY1. Interacts with HGS, NANOG and ZCCHC12 (By similarity). Upon C-terminus phosphorylation: forms trimers with another SMAD1 and the co-SMAD SMAD4. Interacts with PEBP2-alpha subunit, CREB-binding protein (CBP), p300, SMURF1, SMURF2, USP15 and HOXC8. Associates with ZNF423 or ZNF521 in response to BMP2 leading to activate transcription of BMP target genes. Interacts with SKOR1. Interacts (via MH2 domain) with LEMD3. Binding to LEMD3 results in at least a partial reduction of receptor-mediated phosphorylation. Forms a ternary complex with PSMB4 and OAZ1 before PSMB4 is incorporated into the 20S proteasome. Found in a macromolecular complex with FAM83G. Interacts (via MH2 domain) with FAM83G (via MH2 domain); in a SMAD4-independent manner. Interacts with ZC3H3 (By similarity). Interacts with TMEM119 (By similarity). Interacts (via MH1 and MH2 domains) with ZNF8. |
| Research Area: | Stem Cells |
| Subcellular Location: | Cytoplasm Nucleus Cytoplasmic in the absence of ligand. Migrates to the nucleus when complexed with SMAD4 (PubMed:15647271). Co-localizes with LEMD3 at the nucleus inner membrane (PubMed:15647271). |
| Storage: | Please see kit components below for exact storage details |
| Note: | For research use only |
| UniProt Protein Function: | SMAD1: transcription factor phosphorylated and activated by bone morphogenetic protein type 1 receptor kinases. Participates in a wide range of critical processes including morphogenesis, cell-fate determination, proliferation, differentiation and apoptosis. Phosphorylated forms dimerize with collaborating Smad4 and are translocated into the nucleus, where the transcription of target genes is stimulated. |
| UniProt Protein Details: | Protein type:DNA-binding; Transcription factor Chromosomal Location of Human Ortholog: 4q31 Cellular Component: nucleoplasm; transcription factor complex; protein complex; cytoplasm; integral to membrane; nuclear inner membrane; intracellular; cytosol; nucleus Molecular Function:identical protein binding; protein binding; metal ion binding; receptor signaling protein activity; protein kinase binding; transcription factor activity; transforming growth factor beta receptor, pathway-specific cytoplasmic mediator activity Biological Process: wound healing; cardiac muscle cell proliferation; primary microRNA processing; embryonic pattern specification; signal transduction; protein amino acid phosphorylation; mesodermal cell fate commitment; BMP signaling pathway; negative regulation of cell proliferation; homeostatic process; ureteric bud development; transforming growth factor beta receptor signaling pathway; midbrain development; inflammatory response; hindbrain development; response to drug; transcription, DNA-dependent; MAPKKK cascade; positive regulation of dendrite morphogenesis; response to organic nitrogen; SMAD protein complex assembly; positive regulation of osteoblast differentiation; gamete generation; cartilage development; positive regulation of transcription from RNA polymerase II promoter; osteoblast fate commitment |
| NCBI Summary: | The protein encoded by this gene belongs to the SMAD, a family of proteins similar to the gene products of the Drosophila gene 'mothers against decapentaplegic' (Mad) and the C. elegans gene Sma. SMAD proteins are signal transducers and transcriptional modulators that mediate multiple signaling pathways. This protein mediates the signals of the bone morphogenetic proteins (BMPs), which are involved in a range of biological activities including cell growth, apoptosis, morphogenesis, development and immune responses. In response to BMP ligands, this protein can be phosphorylated and activated by the BMP receptor kinase. The phosphorylated form of this protein forms a complex with SMAD4, which is important for its function in the transcription regulation. This protein is a target for SMAD-specific E3 ubiquitin ligases, such as SMURF1 and SMURF2, and undergoes ubiquitination and proteasome-mediated degradation. Alternatively spliced transcript variants encoding the same protein have been observed. [provided by RefSeq, Jul 2008] |
| UniProt Code: | Q15797 |
| NCBI GenInfo Identifier: | 13633915 |
| NCBI Gene ID: | 4086 |
| NCBI Accession: | Q15797.1 |
| UniProt Secondary Accession: | Q15797,Q16636, Q9UFT8, A8KAJ0, D3DNZ9, |
| UniProt Related Accession: | Q15797 |
| Molecular Weight: | 465 |
| NCBI Full Name: | Mothers against decapentaplegic homolog 1 |
| NCBI Synonym Full Names: | SMAD family member 1 |
| NCBI Official Symbol: | SMAD1 |
| NCBI Official Synonym Symbols: | BSP1; JV41; BSP-1; JV4-1; MADH1; MADR1 |
| NCBI Protein Information: | mothers against decapentaplegic homolog 1; MAD homolog 1; Mad-related protein 1; TGF-beta signaling protein 1; mothers against DPP homolog 1; SMAD, mothers against DPP homolog 1; MAD, mothers against decapentaplegic homolog 1; transforming growth factor-beta signaling protein 1; transforming growth factor-beta-signaling protein 1 |
| UniProt Protein Name: | Mothers against decapentaplegic homolog 1 |
| UniProt Synonym Protein Names: | JV4-1; Mad-related protein 1; SMAD family member 1; SMAD 1; Smad1; hSMAD1; Transforming growth factor-beta-signaling protein 1; BSP-1 |
| Protein Family: | Mothers against decapentaplegic |
| UniProt Gene Name: | SMAD1 |
| UniProt Entry Name: | SMAD1_HUMAN |
| 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. |
