Human UBE2C (Ubiquitin Conjugating Enzyme E2C) ELISA Kit (HUES02318)
- Product Type:
- ELISA Kit
- 96 Assays
- ELISA Type:
- Tested Sample Types:
- Serum, plasma and other biological fluids
|Detection Range:||0.16-10 ng/mL|
|Sample Volume Required Per Well:||100µL|
|Sample Type:||Serum, plasma and other biological fluids|
|Specificity:||This kit recognizes Human UBE2C in samples. No significant cross-reactivity or interference between Human UBE2C and analogues was observed.|
This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human UBE2C. Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human UBE2C and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human UBE2C, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Human UBE2C. The concentration of Human UBE2C in samples can be calculated by comparing the OD of the samples to the standard curve.
|UniProt Protein Function:||UBE2C: Accepts ubiquitin from the E1 complex and catalyzes its covalent attachment to other proteins. In vitro catalyzes 'Lys- 11'- and 'Lys-48'-linked polyubiquitination. Acts as an essential factor of the anaphase promoting complex/cyclosome (APC/C), a cell cycle-regulated ubiquitin ligase that controls progression through mitosis. Acts by initiating 'Lys-11'-linked polyubiquitin chains on APC/C substrates, leading to the degradation of APC/C substrates by the proteasome and promoting mitotic exit. Belongs to the ubiquitin-conjugating enzyme family.|
|UniProt Protein Details:|
Protein type:Ligase; Cell cycle regulation; Ubiquitin conjugating system; Ubiquitin ligase; EC 6. 3. 2. 19
Chromosomal Location of Human Ortholog: 20q13. 12
Cellular Component: anaphase-promoting complex; cytoplasm; cytosol; nucleoplasm
Molecular Function:protein binding; ubiquitin protein ligase binding; ubiquitin-protein ligase activity
Biological Process: anaphase-promoting complex-dependent proteasomal ubiquitin-dependent protein catabolic process; cyclin catabolic process; exit from mitosis; negative regulation of ubiquitin-protein ligase activity during mitotic cell cycle; positive regulation of exit from mitosis; positive regulation of ubiquitin-protein ligase activity during mitotic cell cycle; proteasomal ubiquitin-dependent protein catabolic process; protein ubiquitination; protein ubiquitination during ubiquitin-dependent protein catabolic process; regulation of mitotic metaphase/anaphase transition; regulation of ubiquitin-protein ligase activity during mitotic cell cycle; ubiquitin-dependent protein catabolic process
|NCBI Summary:||The modification of proteins with ubiquitin is an important cellular mechanism for targeting abnormal or short-lived proteins for degradation. Ubiquitination involves at least three classes of enzymes: ubiquitin-activating enzymes, ubiquitin-conjugating enzymes, and ubiquitin-protein ligases. This gene encodes a member of the E2 ubiquitin-conjugating enzyme family. The encoded protein is required for the destruction of mitotic cyclins and for cell cycle progression, and may be involved in cancer progression. Multiple transcript variants encoding different isoforms have been found for this gene. Pseudogenes of this gene have been defined on chromosomes 4, 14, 15, 18, and 19. [provided by RefSeq, Aug 2013]|
|NCBI GenInfo Identifier:||3915178|
|NCBI Gene ID:||11065|
|NCBI Accession:||O00762. 1|
|UniProt Secondary Accession:||O00762,A6NP33, E1P5N7, G3XAB7,|
|UniProt Related Accession:||O00762|
|Molecular Weight:||17,373 Da|
|NCBI Full Name:||Ubiquitin-conjugating enzyme E2 C|
|NCBI Synonym Full Names:||ubiquitin conjugating enzyme E2 C|
|NCBI Official Symbol:||UBE2C|
|NCBI Official Synonym Symbols:||UBCH10; dJ447F3. 2|
|NCBI Protein Information:||ubiquitin-conjugating enzyme E2 C|
|UniProt Protein Name:||Ubiquitin-conjugating enzyme E2 C|
|UniProt Synonym Protein Names:||(E3-independent) E2 ubiquitin-conjugating enzyme C (EC:2. 3. 2. 24|
|Protein Family:||Ubiquitin-conjugating enzyme|
|UniProt Gene Name:||UBE2C|
|UniProt Entry Name:||UBE2C_HUMAN|
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e. g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human UBE2C were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human UBE2C were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||6.00||5.04||4.87||5.56||4.61||3.60|
The recovery of Human UBE2C spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
|Sample Type||Range (%)||Average Recovery (%)|
|EDTA plasma (n=5)||89-101||95|
|Cell culture media (n=5)||91-106||97|
Samples were spiked with high concentrations of Human UBE2C and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
|Serum (n=5)||EDTA plasma (n=5)||Cell culture media (n=5)|
An unopened kit can be stored at 4°C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
|Micro ELISA Plate(Dismountable)||8 wells ×12 strips||-20°C, 6 months|
|Reference Standard||2 vials|
|Concentrated Biotinylated Detection Ab (100×)||1 vial, 120 µL|
|Concentrated HRP Conjugate (100×)||1 vial, 120 µL||-20°C(shading light), 6 months|
|Reference Standard & Sample Diluent||1 vial, 20 mL||4°C, 6 months|
|Biotinylated Detection Ab Diluent||1 vial, 14 mL|
|HRP Conjugate Diluent||1 vial, 14 mL|
|Concentrated Wash Buffer (25×)||1 vial, 30 mL|
|Substrate Reagent||1 vial, 10 mL||4°C(shading light)|
|Stop Solution||1 vial, 10 mL||4°C|
|Plate Sealer||5 pieces|
|Product Description||1 copy|
|Certificate of Analysis||1 copy|
- Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- Aliquot 100 µL of standard solutions into the standard wells.
- Add 100 µL of Sample / Standard dilution buffer into the control (zero) well.
- Add 100 µL of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- Cover the plate with the sealer provided in the kit and incubate for 90 min at 37 °C.
- Aspirate the liquid from each well, do not wash. Immediately add 100 µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix. Incubate for 1 hour at 37 °C.
- Aspirate or decant the solution from the plate and add 350 µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- Add 100 µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37 °C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- Add 90 µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37 °C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
- Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.