Mouse Epigenetics and Nuclear Signaling ELISA Kits
Mouse BMI-1(Polycomb complex protein BMI-1) ELISA Kit
- SKU:
- MOFI01238
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- P25916
- Sensitivity:
- 4.688pg/ml
- Range:
- 7.813-500pg/ml
- ELISA Type:
- Sandwich
- Synonyms:
- B lymphoma Mo-MLV insertion region 1 homolog, flvi-2, bmi-1, murine leukemia viral, bmi-1 oncogene homolog, oncogene BMI-1
- Reactivity:
- Mouse
- Research Area:
- Epigenetics and Nuclear Signaling
Description
| Product Name: | Mouse BMI-1(Polycomb complex protein BMI-1) ELISA Kit |
| Product Code: | MOFI01238 |
| Size: | 96 Assays |
| Alias: | B lymphoma Mo-MLV insertion region 1 homolog, flvi-2, bmi-1, murine leukemia viral, bmi-1 oncogene homolog, oncogene BMI-1 |
| Detection Method: | Sandwich ELISA |
| Application: | This immunoassay kit allows for the in vitro quantitative determination of Mouse BMI-1 concentrations in serum plasma and other biological fluids. |
| Sensitivity: | 4.688pg/ml |
| Range: | 7.813-500pg/ml |
| Storage: | 4°C for 6 months |
| Note: | For Research Use Only |
| Recovery: | Matrices listed below were spiked with certain level of Mouse BMI-1 and the recovery rates were calculated by comparing the measured value to the expected amount of Mouse BMI-1 in samples. Not Available. |
| Linearity: | The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Mouse BMI-1 and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected. Not Available. |
| Intra Assay: | CV <8% |
| Inter Assay: | CV <10% |
| Component | Quantity | Storage |
| ELISA Microplate (Dismountable) | 8-12 strips | 4°C for 6 months |
| Lyophilized Standard | 2 | 4°C/-20°C |
| Sample/Standard Dilution Buffer | 20ml | 4°C |
| Biotin-labeled Antibody(Concentrated) | 120ul | 4°C (Protect from light) |
| Antibody Dilution Buffer | 10ml | 4°C |
| HRP-Streptavidin Conjugate(SABC) | 120ul | 4°C (Protect from light) |
| SABC Dilution Buffer | 10ml | 4°C |
| TMB Substrate | 10ml | 4°C (Protect from light) |
| Stop Solution | 10ml | 4°C |
| Wash Buffer(25X) | 30ml | 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
| Uniprot | P25916 |
| UniProt Protein Function: | BMI1: Component of a Polycomb group (PcG) multiprotein PRC1- like complex, a complex class required to maintain the transcriptionally repressive state of many genes, including Hox genes, throughout development. PcG PRC1 complex acts via chromatin remodeling and modification of histones; it mediates monoubiquitination of histone H2A 'Lys-119', rendering chromatin heritably changed in its expressibility. In the PRC1 complex, it is required to stimulate the E3 ubiquitin-protein ligase activity of RNF2/RING2. Component of a PRC1-like complex. Interacts with RING1 and RING2. Interacts vwith CBX7 and CBX8. Interacts with SPOP. Part of a complex consisting of BMI1, CUL3 and SPOP. Interacts with E4F1. |
| UniProt Protein Details: | Protein type:Ligase; EC 6.3.2.-; Ubiquitin conjugating system; Ubiquitin ligase; Motility/polarity/chemotaxis; Oncoprotein Cellular Component: heterochromatin; nuclear body; nucleus; PcG protein complex; ubiquitin ligase complex Molecular Function:chromatin binding; protein binding; sequence-specific DNA binding Biological Process: brain development; cellular process; DNA methylation; embryonic skeletal development; embryonic skeletal morphogenesis; histone acetylation; histone ubiquitination; humoral immune response; in utero embryonic development; negative regulation of transcription from RNA polymerase II promoter; positive regulation of B cell proliferation; positive regulation of immature T cell proliferation in the thymus; rostrocaudal neural tube patterning; skeletal development; somatic stem cell division |
| UniProt Code: | P25916 |
| NCBI GenInfo Identifier: | 115066 |
| NCBI Gene ID: | 12151 |
| NCBI Accession: | P25916.1 |
| UniProt Related Accession: | P25916 |
| Molecular Weight: | 36,708 Da |
| NCBI Full Name: | Polycomb complex protein BMI-1 |
| NCBI Synonym Full Names: | Bmi1 polycomb ring finger oncogene |
| NCBI Official Symbol: | Bmi1Â Â |
| NCBI Official Synonym Symbols: | Bmi-1; Pcgf4; AW546694Â Â |
| NCBI Protein Information: | polycomb complex protein BMI-1 |
| UniProt Protein Name: | Polycomb complex protein BMI-1 |
| UniProt Synonym Protein Names: | Polycomb group RING finger protein 4 |
| Protein Family: | Polycomb complex protein |
| UniProt Gene Name: | Bmi1Â Â |
| UniProt Entry Name: | BMI1_MOUSE |
*Note: Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
| Step | Procedure |
| 1. | Set standard, test sample and control (zero) wells on the pre-coated plate respectively, and then, record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (zero) wells! |
| 2. | Aliquot 0.1ml standard solutions into the standard wells. |
| 3. | Add 0.1 ml of Sample / Standard dilution buffer into the control (zero) well. |
| 4. | Add 0.1 ml of properly diluted sample (Human serum, plasma, tissue homogenates and other biological fluids.) into test sample wells. |
| 5. | Seal the plate with a cover and incubate at 37 °C for 90 min. |
| 6. | Remove the cover and discard the plate content, clap the plate on the absorbent filter papers or other absorbent material. Do NOT let the wells completely dry at any time. Wash plate X2. |
| 7. | Add 0.1 ml of Biotin- detection antibody working solution into the above wells (standard, test sample & zero wells). Add the solution at the bottom of each well without touching the side wall. |
| 8. | Seal the plate with a cover and incubate at 37°C for 60 min. |
| 9. | Remove the cover, and wash plate 3 times with Wash buffer. Let wash buffer rest in wells for 1 min between each wash. |
| 10. | Add 0.1 ml of SABC working solution into each well, cover the plate and incubate at 37°C for 30 min. |
| 11. | Remove the cover and wash plate 5 times with Wash buffer, and each time let the wash buffer stay in the wells for 1-2 min. |
| 12. | Add 90 µL of TMB substrate into each well, cover the plate and incubate at 37°C in dark within 10-20 min. (Note: This incubation time is for reference use only, the optimal time should be determined by end user.) And the shades of blue can be seen in the first 3-4 wells (with most concentrated standard solutions), the other wells show no obvious color. |
| 13. | Add 50 µL of Stop solution into each well and mix thoroughly. The color changes into yellow immediately. |
| 14. | Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution. |
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. |
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