Human CDC23 (Cell Division Cycle Protein 23) ELISA Kit (HUES01837)
- SKU:
- HUES01837
Description
Human CDC23 (Cell Division Cycle Protein 23) ELISA Kit
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 CDC23 . 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 CDC23 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 CDC23, 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 CDC23. The concentration of Human CDC23 in samples can be calculated by comparing the OD of the samples to the standard curve.
| Assay type | Sandwich |
| Format | 96T |
| Assay time | 4.5h |
| Reactivity | Human |
| Detection Method | Colormetric |
| Detection Range | 31.25-2000 pg/mL |
| Sensitivity | 18.75 pg/mL |
| Sample Volume Required Per Well | 100uL |
| Sample Type | Serum, plasma and other biological fluids |
Specificity
This kit recognizes Human CDC23 in samples. No significant cross-reactivity or interference between Human CDC23 and analogues was observed.
Typical Data
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.
| Concentration (pg/mL) | O.D | Average | Corrected |
|---|---|---|---|
| 2000 | 2.382 2.396 | 2.389 | 2.301 |
| 1000 | 1.556 1.584 | 1.57 | 1.482 |
| 500 | 0.935 0.929 | 0.932 | 0.844 |
| 250 | 0.503 0.517 | 0.51 | 0.422 |
| 125 | 0.27 0.246 | 0.258 | 0.17 |
| 62.5 | 0.191 0.187 | 0.189 | 0.101 |
| 31.25 | 0.132 0.148 | 0.14 | 0.052 |
| 0 | 0.086 0.09 | 0.088 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human CDC23 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human CDC23 were tested on 3 different plates, 20 replicates in each plate.
| Intra-assay Precision | Inter-assay Precision | |||||
|---|---|---|---|---|---|---|
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean (pg/mL) | 96.50 | 283.10 | 738.09 | 88.24 | 294.96 | 703.76 |
| Standard deviation | 5.32 | 11.83 | 27.31 | 5.83 | 17.40 | 33.01 |
| C V (%) | 5.51 | 4.18 | 3.70 | 6.61 | 5.90 | 4.69 |
Recovery
The recovery of Human CDC23 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
| Sample Type | Range (%) | Average Recovery (%) |
|---|---|---|
| Serum (n=5) | 90-105 | 97 |
| EDTA plasma (n=5) | 93-109 | 100 |
| Cell culture media (n=5) | 86-98 | 93 |
Linearity
Samples were spiked with high concentrations of Human CDC23 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) | ||
|---|---|---|---|---|
| 1:2 | Range (%) | 94-110 | 85-99 | 90-104 |
| Average (%) | 101 | 91 | 96 | |
| 1:4 | Range (%) | 87-103 | 81-92 | 81-94 |
| Average (%) | 94 | 87 | 88 | |
| 1:8 | Range (%) | 93-108 | 85-98 | 85-97 |
| Average (%) | 99 | 92 | 92 | |
| 1:16 | Range (%) | 92-107 | 85-99 | 84-96 |
| Average (%) | 99 | 90 | 89 |
Kit Components & Storage
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.
| Item | Specifications | Storage |
|---|---|---|
| Micro ELISA Plate(Dismountable) | 8 wells X12 strips | -20'C, 6 months |
| Reference Standard | 2 vials | |
| Concentrated Biotinylated Detection Ab (100X) | 1 vial, 120 uL | |
| Concentrated HRP Conjugate (100X) | 1 vial, 120 uL | -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 (25X) | 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 |
Human CDC23 (Cell Division Cycle Protein 23) ELISA Kit (HUES01837) Assay procedure
- 1. 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.
- 2. Aliquot 100µl of standard solutions into the standard wells.
- 3. Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- 4. Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- 5. Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- 6. Aspirate the liquid from each well, do not wash. Immediately add 100µL 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.
- 7. Aspirate or decant the solution from the plate and add 350µL 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.
- 8. Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- 9. Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- 10. Add 90µL 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.
- 11. Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- 12. Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.
Human CDC23 (Cell Division Cycle Protein 23) ELISA Kit (HUES01837) Protein Information
| UniProt Protein Function: | CDC23: Component of the anaphase promoting complex/cyclosome (APC/C), a cell cycle-regulated E3 ubiquitin ligase that controls progression through mitosis and the G1 phase of the cell cycle. The APC/C complex acts by mediating ubiquitination and subsequent degradation of target proteins: it mainly mediates the formation of 'Lys-11'-linked polyubiquitin chains and, to a lower extent, the formation of 'Lys-48'- and 'Lys-63'-linked polyubiquitin chains. Belongs to the APC8/CDC23 family. 3 isoforms of the human protein are produced by alternative splicing. |
| UniProt Protein Details: | Protein type:Cell cycle regulation Chromosomal Location of Human Ortholog: 5q31 Cellular Component: nucleoplasm; anaphase-promoting complex; intracellular; cytosol Molecular Function:ubiquitin-protein ligase activity Biological Process: positive regulation of ubiquitin-protein ligase activity during mitotic cell cycle; ubiquitin-dependent protein catabolic process; mitosis; negative regulation of ubiquitin-protein ligase activity during mitotic cell cycle; regulation of ubiquitin-protein ligase activity during mitotic cell cycle; anaphase-promoting complex-dependent proteasomal ubiquitin-dependent protein catabolic process; cell division; regulation of mitotic metaphase/anaphase transition; mitotic cell cycle spindle assembly checkpoint; regulation of exit from mitosis; mitotic cell cycle; mitotic metaphase plate congression; mitotic metaphase/anaphase transition |
| NCBI Summary: | The protein encoded by this gene shares strong similarity with Saccharomyces cerevisiae Cdc23, a protein essential for cell cycle progression through the G2/M transition. This protein is a component of anaphase-promoting complex (APC), which is composed of eight protein subunits and highly conserved in eukaryotic cells. APC catalyzes the formation of cyclin B-ubiquitin conjugate that is responsible for the ubiquitin-mediated proteolysis of B-type cyclins. This protein and 3 other members of the APC complex contain the TPR (tetratricopeptide repeat), a protein domain important for protein-protein interaction. [provided by RefSeq, Jul 2008] |
| UniProt Code: | Q9UJX2 |
| NCBI GenInfo Identifier: | 254763423 |
| NCBI Gene ID: | 8697 |
| NCBI Accession: | Q9UJX2.3 |
| UniProt Secondary Accession: | Q9UJX2,O75433, Q53FN2, Q9BS73, A8K6E5, B4E3A2, B7WP05 D3DQB7, |
| UniProt Related Accession: | Q9UJX2 |
| Molecular Weight: | 55,882 Da |
| NCBI Full Name: | Cell division cycle protein 23 homolog |
| NCBI Synonym Full Names: | cell division cycle 23 |
| NCBI Official Symbol: | CDC23 |
| NCBI Official Synonym Symbols: | APC8; CUT23; ANAPC8 |
| NCBI Protein Information: | cell division cycle protein 23 homolog; cyclosome subunit 8; cell division cycle 23 homolog; anaphase promoting complex subunit 8; anaphase-promoting complex subunit 8 |
| UniProt Protein Name: | Cell division cycle protein 23 homolog |
| UniProt Synonym Protein Names: | Anaphase-promoting complex subunit 8; APC8; Cyclosome subunit 8 |
| Protein Family: | Anaphase-promoting complex |
| UniProt Gene Name: | CDC23 |
| UniProt Entry Name: | CDC23_HUMAN |
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