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Boster's ELISA Kit is for the detection of human Hsp70 in cell lysates, and tissue extracts. Each kit contains sufficient components to quantitate the Hsp70 concentration in up to 40 samples, tested in duplicate.
| Product Name | HSP70 ELISA Kit |
|---|---|
| SKU/Catalog Number | EK7106 |
| Description | Colorimetric detection of HSP70. 96wells/kit, with removable strips. |
| Cite This Product | HSP70 ELISA Kit (Boster Biological Technology, Pleasanton CA, USA, Catalog # EK7106) |
| Validated Species | Canine, Goat, Human |
| Application | ELISA *Our Boster Guarantee covers the use of this product in the above tested applications. **For positive and negative control design, consult "Tissue specificity" under Protein Target Info. |
| Cross Reactivity | There is no detectable cross-reactivity. |
| Pack Size | 96wells/kit, with removable strips. |
| Sensitivity | 0.18 ng/ml *Sensitivity, or Lower Limit of Detection (LLD), is the minimum level of target protein the ELISA assay can detect. We measure 20 blank wells and if the O.D. value is 2 standard deviations higher than the blanks' average O.D. the sample can be deemed positive. |
|---|---|
| Assay Range | 0.781 - 50 ng/mL *This assay range is determined using common samples. For samples with low target protein concentrations, users can adjust the standard curve to extend the lower limit of assay range. |
| Sample Type | Cell lysates, Tissue *The above listed samples are the ones valided with the assay. If you do not see your sample of interest listed, as long as there is enough level of target protein present in the sample, this Picokine? ELISA kit should detect it. **For protocol and tips regarding preparing your sample of interest, please check our ELISA sample preparation guide. |
| Storage | Store at 4°C. |
| Description | Quantity |
| Anti-Hsp70 Immunoassay Plate | 1 Plate |
| 5X Hsp70 Extraction Reagent | 1 vial/10 ml |
| Recombinant Hsp70 Standard | 2 vials |
| Standard and Sample Diluent | 1 vial/ 50 ml |
| 10X Wash Buffer Concentrate | 1 vial/100 ml |
| Anti-Hsp70 Biotinylated Antibody Concentrate | 1 vial/150 ?l |
| Anti-Hsp70 Biotinylated Antibody Diluent | 1 vial/ 13 ml |
| Streptavidin: HRP Concentrate | 1 vial/150 ?l |
| Streptavidin: HRP Diluent | 1 vial/ 13 ml |
| TMB Substrate | 1 vial/ 13 ml |
| Stop Solution | 1 vial/ 13 ml |
| Pre-treatment Buffer | 1 vial/ 13 ml |
1. Ultra pure water.
2. Additional reagents and materials for cell lysate and tissue extract preparation, including protease inhibitors.
3. Precision pipettors, with disposable plastic tips.
4. Polypropylene or polyethylene tubes to prepare samples ? do not use polystyrene, polycarbonate or glass tubes.
5. A container to prepare 1X Wash Buffer.
6. A wash bottle or an automated 96-well plate washer.
7. Disposable reagent reservoirs.
8. A standard microtiter plate reader for measuring absorbance at 450 nm.
9. Adhesive plate sealers.
You can check the tissue specificity below for information on selecting positive and negative control.
| Gene Name | HSPA1A/HSPA1B |
|---|---|
| Protein Name | Heat shock 70 kDa protein 1A/1B |
| Protein Function | Molecular chaperone implicated in a wide variety of cellular processes, including protection of the proteome from stress, folding and transport of newly synthesized polypeptides, activation of proteolysis of misfolded proteins and the formation and dissociation of protein complexes. Plays a pivotal role in the protein quality control system, ensuring the correct folding of proteins, the re-folding of misfolded proteins and controlling the targeting of proteins for subsequent degradation. This is achieved through cycles of ATP binding, ATP hydrolysis and ADP release, mediated by co-chaperones. The co-chaperones have been shown to not only regulate different steps of the ATPase cycle, but they also have an individual specificity such that one co-chaperone may promote folding of a substrate while another may promote degradation. The affinity for polypeptides is regulated by its nucleotide bound state. In the ATP-bound form, it has a low affinity for substrate proteins. However, upon hydrolysis of the ATP to ADP, it undergoes a conformational change that increases its affinity for substrate proteins. It goes through repeated cycles of ATP hydrolysis and nucleotide exchange, which permits cycles of substrate binding and release. The co-chaperones are of three types: J-domain co-chaperones such as HSP40s (stimulate ATPase hydrolysis by HSP70), the nucleotide exchange factors (NEF) such as BAG1/2/3 (facilitate conversion of HSP70 from the ADP- bound to the ATP-bound state thereby promoting substrate release), and the TPR domain chaperones such as HOPX and STUB1 (PubMed:24012426, PubMed:26865365, PubMed:24318877). Maintains protein homeostasis during cellular stress through two opposing mechanisms: protein refolding and degradation. Its acetylation/deacetylation state determines whether it functions in protein refolding or protein degradation by controlling the competitive binding of co-chaperones HOPX and STUB1. During the early stress response, the acetylated form binds to HOPX which assists in chaperone-mediated protein refolding, thereafter, it is deacetylated and binds to ubiquitin ligase STUB1 that promotes ubiquitin-mediated protein degradation (PubMed:27708256). Regulates centrosome integrity during mitosis, and is required for the maintenance of a functional mitotic centrosome that supports the assembly of a bipolar mitotic spindle (PubMed:27137183). Enhances STUB1-mediated SMAD3 ubiquitination and degradation and facilitates STUB1-mediated inhibition of TGF-beta signaling (PubMed:24613385). Essential for STUB1-mediated ubiquitination and degradation of FOXP3 in regulatory T-cells (Treg) during inflammation (PubMed:23973223). Negatively regulates heat shock- induced HSF1 transcriptional activity during the attenuation and recovery phase period of the heat shock response (PubMed:9499401). |
| Tissue Specificity | HSPA1B is testis-specific. |
| Subcellular Localization | Cytoplasm. |
| Uniprot ID | P0DMV8/P0DMV9 |
| Alternative Names | Heat shock 70 kDa protein 1A; Heat shock 70 kDa protein 1B |
| Research Areas | Cancer, Heat Shock, Cell Signaling, Chaperone Proteins, Protein Trafficking, Tumor Biomarkers, OxidativeStress| |
*if product is indicated to react with multiple species, protein info is based on the human gene.
HSP70 genes encode abundant heat-inducible 70-kDa HSPs (HSP70s). In most eukaryotes HSP70 genes exist as part of a multigene family. They are found in most cellular compartments of eukaryotes including nuclei, mitochondria, chloroplasts, the endoplasmic reticulum and the cytosol, as well as in bacteria. The genes show a high degree of conservation, having at least 5O% identity. The N-terminal two thirds of HSP70s are more conserved than the C-terminal third. HSP70 binds ATP with high affinity and possesses a weak ATPase activity which can be stimulated by binding to unfolded proteins and synthetic peptides. When HSC70 (constitutively expressed) present in mammalian cells was truncated, ATP binding activity was found to reside in an N-terminal fragment of 44kDa which lacked peptide binding capacity. Polypeptide binding ability therefore resided within the C-terminal half. The structure of this ATP binding domain displays multiple features of nucleotide binding proteins. All HSP70s, regardless of location, bind proteins, particularly unfolded ones. The molecular chaperones of the HSP70 family recognize and bind to nascent polypeptide chains as well as partially folded intermediates of proteins preventing their aggregation and misfolding. The binding of ATP triggers a critical conformational change leading to the release of the bound substrate protein. The universal ability of HSP70s to undergo cycles of binding to and release from hydrophobic stretches of partially unfolded proteins determines their role in a great variety of vital intracellular functions such as protein synthesis, protein folding and oligomerization and protein transport.
Click the images to enlarge.
Typical Standard Curve for the HSP70 ELISA Kit (Enzyme-Linked Immunosorbent Assay) -EK7106 Assay Type: Sandwich ELISA. Detection Method: Colorimetric Assay. Assay Range: 0.781 - 50 ng/mL.
1. Prepare Standard and samples in Standard and Sample Diluent.
2. Add 100 ?L of Standard to appropriate wells.
3. Add 50 ?L of Pre-Treatment Buffer to all sample wells.
4. Add 50 ?L of sample to appropriate wells.
5. Cover plate with Plate Sealer and incubate at 37°C for 2 hours.
6. Wash plate four times with 1X Was
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