Mouse Cell Signalling ELISA Kits 2
Mouse TJP1 (Tight Junction Protein 1) CLIA Kit (MOES00564)
- SKU:
- MOES00564
- Product Type:
- ELISA Kit
- ELISA Type:
- CLIA Kit
- Size:
- 96 Assays
- Sensitivity:
- 9.38pg/mL
- Range:
- 15.63-1000pg/mL
- ELISA Type:
- Sandwich
- Reactivity:
- Mouse
- Sample Type:
- Serum, plasma and other biological fluids
Description
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Mouse |
| Detection method: | Chemiluminescence |
| Detection range: | 15.63-1000 pg/mL |
| Sensitivity: | 9.38 pg/mL |
| Sample volume: | 100µL |
| Sample type: | Serum, plasma and other biological fluids |
| Repeatability: | CV < 15% |
| Specificity: | This kit recognizes Mouse TJP1 in samples. No significant cross-reactivity or interference between Mouse TJP1 and analogues was observed. |
This kit uses Sandwich-CLIA as the method. The micro CLIA plate provided in this kit has been pre-coated with an antibody specific to Mouse TJP1. Standards or samples are added to the appropriate micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Mouse TJP1 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 Mouse TJP1, biotinylated detection antibody and Avidin-HRP conjugate will appear fluorescence. The Relative light unit (RLU) value is measured spectrophotometrically by the Chemiluminescence immunoassay analyzer. The RLU value is positively associated with the concentration of Mouse TJP1. The concentration of Mouse TJP1 in the samples can be calculated by comparing the RLU of the samples to the standard curve.
| UniProt Protein Function: | Function: The N-terminal may be involved in transducing a signal required for tight junction assembly, while the C-terminal may have specific properties of tight junctions. The alpha domain might be involved in stabilizing junctions. Plays a role in the regulation of cell migration by targeting CDC42BPB to the leading edge of migrating cells |
| UniProt Protein Details: | By similarity. Subunit structure: Homodimer, and heterodimer with TJP2/ZO-2 and TJP3/ZO-3. Interacts with CGN/cingulin, CALM, CXADR, GJD3, HSPA4 and UBN1. Interacts (via ZU5 domain) with CDC42BPB. Interacts (via PDZ domain) with GJA1 By similarity. Interacts with OCLN, claudins, GJA12 and KIRREL1. Interacts with BVES (via the C-terminus cytoplasmic tail). Interacts (via ZU5 domain) with MYZAP. Interacts (via PDZ domains) with ANKRD2 By similarity. Ref. 3 Ref. 4 Ref. 5 Ref. 12 Subcellular location: Cell membrane; Peripheral membrane protein; Cytoplasmic side By similarity. Cell junction tight junction By similarity. Cell junction gap junction. Cytoplasm myofibril sarcomere I band. Note: Moves from the cytoplasm to the cell membrane concurrently with cell-cell contact. Detected at the leading edge of migrating and wounded cells By similarity. Ref. 12 Domain: The second PDZ domain mediates interaction with GJA12. Ref. 4 Post-translational modification: Phosphorylated. Dephosphorylated by PTPRJ By similarity. Sequence similarities: Belongs to the MAGUK family. Contains 1 guanylate kinase-like domain. Contains 3 PDZ (DHR) domains. Contains 1 SH3 domain. Contains 1 ZU5 domain. |
| UniProt Code: | P39447 |
| NCBI GenInfo Identifier: | 303710 |
| NCBI Gene ID: | 21872 |
| NCBI Accession: | BAA03274. 1 |
| UniProt Secondary Accession: | P39447,E9QK00, |
| UniProt Related Accession: | P39447 |
| Molecular Weight: | |
| NCBI Full Name: | ZO-1 |
| NCBI Synonym Full Names: | tight junction protein 1 |
| NCBI Official Symbol: | Tjp1 |
| NCBI Official Synonym Symbols: | ZO1 |
| NCBI Protein Information: | tight junction protein ZO-1; zona occludens protein 1; zonula occludens protein 1 |
| UniProt Protein Name: | Tight junction protein ZO-1 |
| UniProt Synonym Protein Names: | Tight junction protein 1; Zona occludens protein 1; Zonula occludens protein 1 |
| UniProt Gene Name: | Tjp1 |
| UniProt Entry Name: | ZO1_MOUSE |
As the RLU 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) | RLU | Average | Corrected |
| 1000 | 50621 56729 | 53675 | 53650 |
| 500 | 21765 23213 | 22489 | 22464 |
| 250 | 10946 9476 | 10211 | 10186 |
| 125 | 4828 4974 | 4901 | 4876 |
| 62.5 | 2640 2266 | 2453 | 2428 |
| 31.25 | 1336 1226 | 1281 | 1256 |
| 15.63 | 689 727 | 708 | 683 |
| 0 | 24 26 | 25 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Mouse TJP1 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Mouse TJP1 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) | 47.59 | 85.25 | 411.05 | 50.64 | 92.47 | 375.70 |
| Standard deviation | 5.89 | 7.77 | 30.09 | 6.25 | 8.83 | 33.85 |
| C V (%) | 12.38 | 9.11 | 7.32 | 12.34 | 9.55 | 9.01 |
Recovery
The recovery of Mouse TJP1 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) | 88-103 | 94 |
| EDTA plasma (n=5) | 89-105 | 96 |
| Cell culture media (n=5) | 94-110 | 100 |
Linearity
Samples were spiked with high concentrations of Mouse TJP1 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 (%) | 93-107 | 86-101 | 84-98 |
| Average (%) | 99 | 93 | 90 | |
| 1:4 | Range (%) | 96-110 | 88-100 | 88-100 |
| Average (%) | 103 | 95 | 93 | |
| 1:8 | Range (%) | 100-113 | 94-108 | 99-114 |
| Average (%) | 106 | 101 | 107 | |
| 1:16 | Range (%) | 87-98 | 88-98 | 101-116 |
| Average (%) | 93 | 93 | 109 |
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 CLIA 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 A | 1 vial, 5 mL | 4°C (shading light) |
| Substrate Reagent B | 1 vial, 5 mL | 4°C (shading light) |
| 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 100µl of standard solutions into the standard wells.
- Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- Add 100µl 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 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.
- 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.
- Add 100µL 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 100µL of Substrate mixture solution to each well. Cover with a new plate seal andincubate for no more than 5 min at 37°C. Protect the plate from light.
- Determine the RLU value of each well immediately.
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