Rat X-box-binding protein 1 (Xbp1) ELISA Kit
- SKU:
- RTEB1638
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- Q9R1S4
- ELISA Type:
- Sandwich
- Reactivity:
- Rat
Frequently bought together:
Description
| 商品名: | Rat X-box-binding protein 1 (Xbp1) ELISA Kit |
| 製品コード: | RTEB1638 |
| エイリアス: | X-box-binding protein 1, Hepatocarcinogenesis-related transcription factor, HTF, Xbp1, Htf |
| Uniprot: | Q9R1S4 |
| 反応性: | Rat |
| 範囲: | Please contact us for more information |
| 検出方法: | Sandwich |
| サイズ: | 96 Assay |
| 保管所: | Please see kit components below for exact storage details |
| ノート: | For research use only |
| UniProt Protein Function: | XBP1: a transcription factor essential for hepatocyte growth, the differentiation of plasma cells, immunoglobulin secretion, and the unfolded protein response (UPR). XBP1 mRNA is spliced by IRE1 during the UPR to generate a new C-terminus, converting it into a potent unfolded-protein response transcriptional activator and triggering growth arrest and apoptosis. Only the spliced form of XBP1 can activate the UPR efficiently. Activates UPR target genes via direct binding to the UPR element (UPRE). Binds DNA preferably to the CRE-like element 5'-GATGACGTG[TG]N(3)[AT]T-3', and also to some TPA response elements (TRE). Binds to the HLA DR-alpha promoter. Binds to the Tax-responsive element (TRE) of HTLV-I. Up-regulated by ATF6 via direct binding to the ERSE in response to endoplasmic reticulum stress. Genetic variations in XBP1 could be associated with susceptibility to major affective disorder type 7 (MAFD7). Major affective disorders represent a class of mental disorders characterized by a disturbance in mood as their predominant feature. Two human isoforms are produced by alternative splicing. Isoform 1 is also known as XBP-1U. Isoform 2, also known as XBP-1S, is produced by IRE1 in response to endoplasmic reticulum stress. IRE1 cleaves a 26-bp fragment causing a frameshift of the mRNA transcript.Protein type: Transcription factor; DNA-bindingChromosomal Location of Human Ortholog: 22q12.1|22q12Cellular Component: nucleoplasm; endoplasmic reticulum membrane; endoplasmic reticulum; cytoplasm; integral to membrane; integral to endoplasmic reticulum membrane; cytosol; nucleusMolecular Function: protein binding; protein homodimerization activity; DNA binding; protease binding; chromatin DNA binding; protein heterodimerization activity; ubiquitin protein ligase binding; estrogen receptor binding; transcription factor activity; protein kinase bindingBiological Process: ubiquitin-dependent protein catabolic process; transcription from RNA polymerase II promoter; muscle development; phosphoinositide 3-kinase cascade; apoptosis; positive regulation of transcription of target genes involved in unfolded protein response; exocrine pancreas development; regulation of protein stability; negative regulation of transcription from RNA polymerase II promoter; cellular response to glucose starvation; protein transport; serotonin secretion, neurotransmission; positive regulation of MHC class II biosynthetic process; positive regulation of transcription factor import into nucleus; angiogenesis; cell growth; response to electrical stimulus; positive regulation of TOR signaling pathway; positive regulation of autophagy; fatty acid biosynthetic process; response to drug; positive regulation of histone methylation; protein destabilization; unfolded protein response; organelle organization and biogenesis; cellular response to nutrient; positive regulation of immunoglobulin secretion; liver development; positive regulation of immunoglobulin production; cholesterol homeostasis; unfolded protein response, activation of signaling protein activity; cellular protein metabolic process; cellular response to insulin stimulus; positive regulation of T cell differentiation; fatty acid homeostasis; endothelial cell proliferation; neuron development; positive regulation of fat cell differentiation; autophagy; positive regulation of B cell differentiation; positive regulation of transcription from RNA polymerase II promoter; immune response; positive regulation of protein amino acid phosphorylation; sterol homeostasis; vascular endothelial growth factor receptor signaling pathway; negative regulation of apoptosisDisease: Major Affective Disorder 7 |
| UniProt Protein Details: | |
| NCBI Summary: | This gene encodes a transcription factor that regulates MHC class II genes by binding to a promoter element referred to as an X box. This gene product is a bZIP protein, which was also identified as a cellular transcription factor that binds to an enhancer in the promoter of the T cell leukemia virus type 1 promoter. It may increase expression of viral proteins by acting as the DNA binding partner of a viral transactivator. It has been found that upon accumulation of unfolded proteins in the endoplasmic reticulum (ER), the mRNA of this gene is processed to an active form by an unconventional splicing mechanism that is mediated by the endonuclease inositol-requiring enzyme 1 (IRE1). The resulting loss of 26 nt from the spliced mRNA causes a frame-shift and an isoform XBP1(S), which is the functionally active transcription factor. The isoform encoded by the unspliced mRNA, XBP1(U), is constitutively expressed, and thought to function as a negative feedback regulator of XBP1(S), which shuts off transcription of target genes during the recovery phase of ER stress. A pseudogene of XBP1 has been identified and localized to chromosome 5. [provided by RefSeq, Jul 2008] |
| UniProt Code: | Q9R1S4 |
| NCBI GenInfo Identifier: | 47678753 |
| NCBI Gene ID: | 7494 |
| NCBI Accession: | CAG30497.1 |
| UniProt Secondary Accession: | Q9R1S4,O35426, Q9R1S4 |
| UniProt Related Accession: | Q9R1S4,P17861 |
| Molecular Weight: | 40kDa |
| NCBI Full Name: | XBP1 |
| NCBI Synonym Full Names: | X-box binding protein 1 |
| NCBI Official Symbol: | XBP1 |
| NCBI Official Synonym Symbols: | XBP2; TREB5; XBP-1; TREB-5 |
| NCBI Protein Information: | X-box-binding protein 1 |
| UniProt Protein Name: | X-box-binding protein 1 |
| UniProt Synonym Protein Names: | Tax-responsive element-binding protein 5 |
| Protein Family: | X-box-binding protein |
| UniProt Gene Name: | XBP1 |
| UniProt Entry Name: | XBP1_HUMAN |
| 成分 | 額 | 保管所 |
| 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 | - |
その他の必要な材料と設備：
- 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
*ノート: プロトコルは、各バッチ/ロットに固有です。正しい手順については、キットに含まれているプロトコルに従ってください。
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.
| ステップ | プロトコル |
| 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. |
ELISAアッセイを実施する場合、可能な限り最良の結果を達成するためにサンプルを準備することが重要です。以下に、さまざまなサンプルタイプのサンプルを準備するための手順のリストを示します。
| サンプルタイプ: | プロトコル: |
| 血清: | 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. |
| プラズマ: | 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. |
| 尿および脳脊髄液: | 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. |
| 細胞培養上清: | 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. |
| 細胞溶解物: | 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. |
| 組織ホモジネート: | 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. |
| 組織溶解物: | 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. |
| 母乳: | 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. |