Description
Human BIN1/Myc box-dependent-interacting protein 1 ELISA Kit
The Human BIN1 (Myc Box-dependent Interacting Protein 1) ELISA Kit is specifically designed for the accurate detection of BIN1 levels in human serum, plasma, and cell culture supernatants. This kit offers high sensitivity and specificity, ensuring reliable and reproducible results for a variety of research applications.BIN1 is a key protein that interacts with Myc Box, playing a crucial role in cell proliferation and signaling pathways.
Dysregulation of BIN1 has been implicated in various diseases, including cancer, Alzheimer's, and cardiovascular diseases, making it a valuable biomarker for studying these conditions and exploring potential therapeutic interventions.Overall, the Human BIN1 ELISA Kit provides researchers with a powerful tool to analyze BIN1 levels and further understand its role in disease development and progression.
| Product Name: | Human BIN1/Myc box-dependent-interacting protein 1 ELISA Kit |
| Product Code: | HUFI02068 |
| Size: | 96 Assays |
| Alias: | BIN1, Bridging integRator 1, Box-dependent myc-interacting protein 1, Amphiphysin II, Amphiphysin-like protein |
| Detection method: | Sandwich ELISA, Double Antibody |
| Application: | This immunoassay kit allows for the in vitro quantitative determination of Human BIN1 concentrations in serum plasma and other biological fluids. |
| Sensitivity: | 9.375pg/ml |
| Range: | 15.625-1000pg/ml |
| Storage: | 4°C for 6 months |
| Note: | For Research Use Only |
| Recovery: | Matrices listed below were spiked with certain level of Human BIN1 and the recovery rates were calculated by comparing the measured value to the expected amount of Human BIN1 in samples. | ||||||||||||||||
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| Linearity: | The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human BIN1 and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected. | ||||||||||||||||
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| CV(%): | 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 | O00499 |
| UniProt Protein Function: | BIN1: May be involved in regulation of synaptic vesicle endocytosis. May act as a tumor suppressor and inhibits malignant cell transformation. Defects in BIN1 are the cause of centronuclear myopathy type 2 (CNM2). A congenital muscle disorder characterized by progressive muscular weakness and wasting involving mainly limb girdle, trunk, and neck muscles. It may also affect distal muscles. Weakness may be present during childhood or adolescence or may not become evident until the third decade of life. Ptosis is a frequent clinical feature. The most prominent histopathologic features include high frequency of centrally located nuclei in muscle fibers not secondary to regeneration, radial arrangement of sarcoplasmic strands around the central nuclei, and predominance and hypotrophy of type 1 fibers. 11 isoforms of the human protein are produced by alternative splicing. |
| UniProt Protein Details: | Protein type:Tumor suppressor; Cytoskeletal; Adaptor/scaffold; Vesicle Chromosomal Location of Human Ortholog: 2q14 Cellular Component: I band; synaptic vesicle; membrane; axon; T-tubule; cytoplasm; nerve terminal; Z disc; nucleus; actin cytoskeleton Molecular Function:identical protein binding; protein binding; GTPase binding; protein heterodimerization activity; protein complex binding; tau protein binding Biological Process: cell proliferation; muscle cell differentiation; viral reproduction; regulation of neuron differentiation; positive regulation of apoptosis; positive regulation of endocytosis; endocytosis; positive regulation of GTPase activity; positive regulation of astrocyte differentiation Disease: Myopathy, Centronuclear, 2 |
| NCBI Summary: | This gene encodes several isoforms of a nucleocytoplasmic adaptor protein, one of which was initially identified as a MYC-interacting protein with features of a tumor suppressor. Isoforms that are expressed in the central nervous system may be involved in synaptic vesicle endocytosis and may interact with dynamin, synaptojanin, endophilin, and clathrin. Isoforms that are expressed in muscle and ubiquitously expressed isoforms localize to the cytoplasm and nucleus and activate a caspase-independent apoptotic process. Studies in mouse suggest that this gene plays an important role in cardiac muscle development. Alternate splicing of the gene results in ten transcript variants encoding different isoforms. Aberrant splice variants expressed in tumor cell lines have also been described. [provided by RefSeq, Sep 2011] |
| UniProt Code: | O00499 |
| NCBI GenInfo Identifier: | 14916535 |
| NCBI Gene ID: | 274 |
| NCBI Accession: | O00499.1 |
| UniProt Secondary Accession: | O00499,O00297, O00545, O43867, O60552, O60553, O60554 O60555, O75514, O75515, O75516, O75517, |
| UniProt Related Accession: | O00499 |
| Molecular Weight: | 593 |
| NCBI Full Name: | Myc box-dependent-interacting protein 1 |
| NCBI Synonym Full Names: | bridging integrator 1 |
| NCBI Official Symbol: | BIN1 |
| NCBI Official Synonym Symbols: | AMPH2; AMPHL; SH3P9 |
| NCBI Protein Information: | myc box-dependent-interacting protein 1; amphiphysin II; amphiphysin-like protein; box dependant MYC interacting protein 1; box-dependent myc-interacting protein 1 |
| UniProt Protein Name: | Myc box-dependent-interacting protein 1 |
| UniProt Synonym Protein Names: | Amphiphysin II; Amphiphysin-like protein; Box-dependent myc-interacting protein 1; Bridging integrator 1 |
| Protein Family: | Myc box-dependent-interacting protein |
| UniProt Gene Name: | BIN1 |
| UniProt Entry Name: | BIN1_HUMAN |
*Note: Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
Before adding to wells, equilibrate the SABC working solution and TMB substrate for at least 30 min at 37°C. When diluting samples and reagents, they must be mixed completely and evenly. It is recommended to plot a standard curve for each test.
| Step | Protocol |
| 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. |
