Description
Human TSG101 (Tumor susceptibility gene 101 protein) ELISA Kit
The Human TSG101 (Tumor Susceptibility Gene 101) Protein ELISA Kit is a powerful tool for the accurate measurement of TSG101 levels in human samples. This kit offers high sensitivity and specificity, providing dependable and consistent results for a variety of research endeavors.TSG101 is a key protein implicated in tumor formation and progression, playing a role in various cellular processes including cell proliferation and migration. Abnormalities in TSG101 expression have been linked to cancer development, making it a valuable biomarker for understanding the disease and exploring therapeutic interventions.
With the Human TSG101 Protein ELISA Kit, researchers can efficiently analyze TSG101 levels in serum, plasma, or cell culture supernatants, paving the way for insightful discoveries and advancements in cancer research. Don't miss out on this essential tool for studying TSG101 and its implications in oncology.
| Product Name: | Human TSG101 (Tumor susceptibility gene 101 protein) ELISA Kit |
| Product Code: | HUFI05926 |
| Size: | 96 Assays |
| Alias: | Tumor susceptibility gene 101 protein ELISA Kit, ESCRT-I complex subunit TSG101 ELISA Kit, TSG101 ELISA Kit |
| Detection method: | Sandwich ELISA, Double Antibody |
| Application: | This immunoassay kit allows for the in vitro quantitative determination of Human TSG101 (Tumor susceptibility gene 101 protein) concentrations in serum plasma and other biological fluids. |
| Sensitivity: | < 18.75pg/ml |
| Range: | 31.25-2000pg/ml |
| Storage: | 4°C for 6 months |
| Note: | For Research Use Only |
| Recovery: | Matrices listed below were spiked with certain level of Human TSG101 (Tumor susceptibility gene 101 protein) and the recovery rates were calculated by comparing the measured value to the expected amount of Human TSG101 (Tumor susceptibility gene 101 protein) in samples. | ||||||||||||||||
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| Linearity: | The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human TSG101 (Tumor susceptibility gene 101 protein) 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 Protein Function: | TSG101: Component of the ESCRT-I complex, a regulator of vesicular trafficking process. Binds to ubiquitinated cargo proteins and is required for the sorting of endocytic ubiquitinated cargos into multivesicular bodies (MVBs). Mediates the association between the ESCRT-0 and ESCRT-I complex. Required for completion of cytokinesis; the function requires CEP55. May be involved in cell growth and differentiation. Acts as a negative growth regulator. Involved in the budding of many viruses through an interaction with viral proteins that contain a late-budding motif P-[ST]-A-P. This interaction is essential for viral particle budding of numerous retroviruses. Belongs to the ubiquitin-conjugating enzyme family. UEV subfamily. 2 isoforms of the human protein are produced by alternative splicing. |
| UniProt Protein Details: | Protein type:Nuclear receptor co-regulator; Cell cycle regulation Chromosomal Location of Human Ortholog: 11p15 Cellular Component: multivesicular body; late endosome membrane; early endosome; cytoplasm; late endosome; plasma membrane; nucleolus; endosome membrane Molecular Function:ubiquitin binding; protein binding; protein homodimerization activity; DNA binding; ubiquitin protein ligase binding; transcription corepressor activity; calcium-dependent protein binding Biological Process: viral reproduction; ubiquitin-dependent protein catabolic process via the multivesicular body pathway; non-lytic virus budding; protein modification process; viral infectious cycle; virus assembly; endosome transport; keratinocyte differentiation; negative regulation of cell proliferation; protein transport; cell division; viral protein processing; regulation of cell growth; cell cycle arrest; negative regulation of transcription, DNA-dependent Disease: Breast Cancer |
| NCBI Summary: | The protein encoded by this gene belongs to a group of apparently inactive homologs of ubiquitin-conjugating enzymes. The gene product contains a coiled-coil domain that interacts with stathmin, a cytosolic phosphoprotein implicated in tumorigenesis. The protein may play a role in cell growth and differentiation and act as a negative growth regulator. In vitro steady-state expression of this tumor susceptibility gene appears to be important for maintenance of genomic stability and cell cycle regulation. Mutations and alternative splicing in this gene occur in high frequency in breast cancer and suggest that defects occur during breast cancer tumorigenesis and/or progression. [provided by RefSeq, Jul 2008] |
| UniProt Code: | Q99816 |
| NCBI GenInfo Identifier: | 5454140 |
| NCBI Gene ID: | 7251 |
| NCBI Accession: | NP_006283.1 |
| UniProt Secondary Accession: | Q99816,Q9BUM5, |
| UniProt Related Accession: | Q99816 |
| Molecular Weight: | 43,944 Da |
| NCBI Full Name: | tumor susceptibility gene 101 protein |
| NCBI Synonym Full Names: | tumor susceptibility gene 101 |
| NCBI Official Symbol: | TSG101 |
| NCBI Official Synonym Symbols: | TSG10; VPS23 |
| NCBI Protein Information: | tumor susceptibility gene 101 protein; tumor susceptibility protein; ESCRT-I complex subunit TSG101 |
| UniProt Protein Name: | Tumor susceptibility gene 101 protein |
| UniProt Synonym Protein Names: | ESCRT-I complex subunit TSG101 |
| Protein Family: | Tumor susceptibility gene 101 protein |
| UniProt Gene Name: | TSG101 |
| UniProt Entry Name: | TS101_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. |
