Mouse Epigenetics and Nuclear Signaling ELISA Kits
Mouse HSF1 (Heat Shock Factor 1) CLIA Kit (MOES00326)
- SKU:
- MOES00326
- Product Type:
- ELISA Kit
- ELISA Type:
- CLIA Kit
- Size:
- 96 Assays
- Sensitivity:
- 0.09ng/mL
- Range:
- 0.16-10ng/mL
- ELISA Type:
- Sandwich
- Reactivity:
- Mouse
- Sample Type:
- Serum, plasma and other biological fluids
- Research Area:
- Epigenetics and Nuclear Signaling
Description
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Mouse |
| Detection method: | Chemiluminescence |
| Detection range: | 0.16-10 ng/mL |
| Sensitivity: | 0.09 ng/mL |
| Sample volume: | 100µL |
| Sample type: | Serum, plasma and other biological fluids |
| Repeatability: | CV < 15% |
| Specificity: | This kit recognizes Mouse HSF1 in samples. No significant cross-reactivity or interference between Mouse HSF1 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 HSF1. 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 HSF1 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 HSF1, 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 HSF1. The concentration of Mouse HSF1 in the samples can be calculated by comparing the RLU of the samples to the standard curve.
| UniProt Protein Function: | HSF1: a transcription factor that specifically binds heat shock promoter elements (HSE) and activates transcription. Induced in response to heat, heavy metals, and oxidative stress. In higher eukaryotes, HSF is unable to bind to HSEs unless the cells are stressed. Becomes phosphorylated in response to stress, forming homotrimers that bind DNA and activate transcription. Phosphorylation by PLK1 enhances nuclear translocation, and phosphorylation by CaMKII enhances transactivation. Phosphorylation by GSK3 and ERK1 induces binding by 14-3-3 and sequestration in the cytoplasm. In addition, during attenuation from the heat shock response, HSF1 is repressed by direct binding of Hsp70, HSP40, and HSF binding protein 1 (HSBP1). Four alternatively spliced isoforms have been described. |
| UniProt Protein Details: | Protein type:Transcription factor; DNA-binding Cellular Component: nucleoplasm; protein complex; pronucleus; cytoplasm; nucleus Molecular Function:DNA binding; sequence-specific DNA binding; chromatin binding; transcription factor activity Biological Process: transcription from RNA polymerase II promoter; embryonic placenta development; transcription, DNA-dependent; in utero embryonic development; embryonic process involved in female pregnancy; female meiosis; defense response; response to lipopolysaccharide; negative regulation of transcription from RNA polymerase II promoter; positive regulation of multicellular organism growth; negative regulation of tumor necrosis factor production; protein amino acid phosphorylation; negative regulation of cell proliferation; regulation of transcription, DNA-dependent; response to heat; mRNA transcription; spermatogenesis; positive regulation of transcription from RNA polymerase II promoter |
| UniProt Code: | P38532 |
| NCBI GenInfo Identifier: | 33859480 |
| NCBI Gene ID: | 15499 |
| NCBI Accession: | NP_032322. 1 |
| UniProt Secondary Accession: | P38532,O70462, |
| UniProt Related Accession: | P38532 |
| Molecular Weight: | 54,931 Da |
| NCBI Full Name: | heat shock factor protein 1 |
| NCBI Synonym Full Names: | heat shock factor 1 |
| NCBI Official Symbol: | Hsf1 |
| NCBI Official Synonym Symbols: | AA960185; Hsf1beta; Hsf1alpha |
| NCBI Protein Information: | heat shock factor protein 1; HSF 1; HSTF 1; Hsf1gammabeta; Hsf1gammaalpha; heat shock transcription factor 1 beta isoform; heat shock transcription factor 1 alpha isoform; heat shock transcription factor 1 gammabeta isoform; heat shock transcription factor 1 gammaalpha isoform |
| UniProt Protein Name: | Heat shock factor protein 1 |
| UniProt Synonym Protein Names: | Heat shock transcription factor 1; HSTF 1 |
| Protein Family: | Heat shock factor protein |
| UniProt Gene Name: | Hsf1 |
| UniProt Entry Name: | HSF1_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 (ng/mL) | RLU | Average | Corrected |
| 10 | 30219 34723 | 32471 | 32447 |
| 5 | 15291 15649 | 15470 | 15446 |
| 2.5 | 7809 7197 | 7503 | 7479 |
| 1.25 | 3469 3837 | 3653 | 3629 |
| 0.63 | 1797 1725 | 1761 | 1737 |
| 0.31 | 866 782 | 824 | 800 |
| 0.16 | 344 370 | 357 | 333 |
| 0 | 23 25 | 24 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Mouse HSF1 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Mouse HSF1 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 (ng/mL) | 0.50 | 1.46 | 4.90 | 0.46 | 1.50 | 4.74 |
| Standard deviation | 0.05 | 0.16 | 0.52 | 0.05 | 0.16 | 0.48 |
| C V (%) | 10.00 | 10.96 | 10.61 | 10.87 | 10.67 | 10.13 |
Recovery
The recovery of Mouse HSF1 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) | 89-104 | 96 |
| EDTA plasma (n=5) | 90-101 | 95 |
| Cell culture media (n=5) | 94-107 | 99 |
Linearity
Samples were spiked with high concentrations of Mouse HSF1 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 (%) | 99-114 | 100-112 | 94-108 |
| Average (%) | 105 | 106 | 100 | |
| 1:4 | Range (%) | 88-102 | 90-103 | 84-95 |
| Average (%) | 95 | 95 | 90 | |
| 1:8 | Range (%) | 84-95 | 102-119 | 101-112 |
| Average (%) | 90 | 109 | 107 | |
| 1:16 | Range (%) | 89-102 | 90-104 | 91-106 |
| Average (%) | 96 | 96 | 97 |
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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