The Rat ANNEXIN A1 (ANXA1) ELISA Kit is specifically designed for the accurate detection of rat annexin A1 levels in rat serum, plasma, and cell culture supernatants. This kit offers high sensitivity and specificity, ensuring precise and reliable results for a variety of research applications.Annexin A1 is a key protein known for its anti-inflammatory and anti-proliferative properties, playing a crucial role in regulating immune responses and inflammation. Dysregulation of annexin A1 has been linked to various diseases, such as inflammatory disorders and cancer, making it a valuable biomarker for studying these conditions and potential therapeutic interventions.
With the Rat ANNEXIN A1 (ANXA1) ELISA Kit, researchers can accurately measure annexin A1 levels in rat samples, allowing for in-depth investigations into its role in various physiological and pathological processes. This kit provides a valuable tool for advancing scientific research and uncovering new insights into annexin A1 biology.
Product Name:
Rat Annexin A1 (Anxa1) ELISA Kit
SKU:
RTEB0226
Size:
96T
Target:
Rat Annexin A1 (Anxa1)
Synonyms:
Annexin I, Annexin-1, Calpactin II, Calpactin-2, Chromobindin-9, Lipocortin I, Phospholipase A2 inhibitory protein, p35, Anx1
Assay Type:
Sandwich
Detection Method:
ELISA
Reactivity:
Rat
Detection Range:
0.156-10ng/mL
Sensitivity:
0.078ng/mL
Intra CV:
6.4%
Inter CV:
8.7%
Linearity:
Sample
1:2
1:4
1:8
1:16
Serum(N=5)
91-100%
84-94%
111-120%
108-117%
EDTA Plasma(N=5)
104-113%
82-95%
86-98%
88-97%
Heparin Plasma(N=5)
91-101%
90-100%
114-124%
104-117%
Recovery:
Sample Type
Average(%)
Recovery Range(%)
Serum
95
89-101
Plasma
97
91-103
Function:
Plays important roles in the innate immune response as effector of glucocorticoid-mediated responses and regulator of the inflammatory process. Has anti-inflammatory activity. Plays a role in glucocorticoid-mediated down-regulation of the early phase of the inflammatory response. Promotes resolution of inflammation and wound healing (By similarity). Functions at least in part by activating the formyl peptide receptors and downstream signaling cascades. Promotes chemotaxis of granulocytes and monocytes via activation ofthe formyl peptide receptors (By similarity). Contributes to the adaptive immune response by enhancing signaling cascades that are triggered by T-cell activation, regulates differentiation and proliferation of activated T-cells. Promotes the differentiation of T-cells into Th1 cells and negatively regulates differentiation into Th2 cells (By similarity). Has no effect on unstimulated T-cells. Promotes rearrangement of the actin cytoskeleton, cell polarization and cell migration. Negatively regulates hormone exocytosis via activation of the formyl peptide receptors and reorganization of the actin cytoskeleton (By similarity). Has high affinity for Ca(2+) and can bind up to eight Ca(2+) ions (By similarity). Displays Ca(2+)-dependent binding to phospholipid membranes (PubMed:3020049). Plays a role in the formation of phagocytic cups and phagosomes. Plays a role in phagocytosis by mediating the Ca(2+)-dependent interaction between phagosomes and the actin cytoskeleton.
Uniprot:
P07150
Sample Type:
Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids
Specificity:
Natural and recombinant rat Annexin A1
Sub Unit:
Homodimer; non-covalently linked (By similarity). Homodimer; linked by transglutamylation. Homodimers linked by transglutamylation are observed in placenta, but not in other tissues. Interacts with S100A11. Heterotetramer, formed by two molecules each of S100A11 and ANXA1 (By similarity). Interacts with DYSF (By similarity). Interacts with EGFR.
Research Area:
Cardiovascular
Subcellular Location:
Nucleus Cytoplasm Cell projection Cilium Basolateral cell membrane Lateral cell membrane Cell membrane Peripheral membrane protein Apical cell membrane Membrane Peripheral membrane protein Endosome membrane Peripheral membrane protein Secreted Secreted Extracellular space Cell membrane Peripheral membrane protein Extracellular side Early endosome Cytoplasmic vesicle membrane Peripheral membrane protein Secreted Exosome Cytoplasmic vesicle Secretory vesicle lumen Cell projection Phagocytic cup Colocalizes with actin fibers at phagocytic cups. Secreted, at least in part via exosomes and other secretory vesicles. Detected in exosomes and other extracellular vesicles. Secretion is increased in response to wounding and inflammation (By similarity). Detected in gelatinase granules in resting neutrophils. Neutrophil adhesion to endothelial cells stimulates secretion via gelatinase granules, but foreign particle phagocytosis has no effect. Displays calcium-dependent binding to phospholipid membranes (PubMed:3020049).
Storage:
Please see kit components below for exact storage details
Note:
For research use only
UniProt Protein Function:
ANXA1: a calcium/phospholipid-binding protein with which promotes membrane fusion and is involved in endocytosis. Has anti-inflammatory properties and inhibits phospholipase A2 activity. Accumulates on internalized vesicles after EGF-stimulated endocytosis, suggesting that it may be required for a late stage in inward vesiculation. Phosphorylated by PKC, EGFR and Chak1. Phosphorylation results in loss of the inhibitory activity. Annexins are a family of structurally related proteins whose common property is calcium-dependent binding to phospholipids. There are at least ten different annexins in mammalian species. Annexins do not contain signal peptides, yet some annexins (A1, A2 and A5) appear to be secreted in a physiologically regulated fashion.Protein type: Calcium-binding; Lipid-bindingChromosomal Location of Human Ortholog: 9q21.13Cellular Component: extracellular space; protein complex; focal adhesion; cell surface; mast cell granule; basolateral plasma membrane; extracellular region; cornified envelope; cilium; cytoplasm; mitochondrial membrane; plasma membrane; nucleus; vesicle; endosome; sarcolemmaMolecular Function: protein binding, bridging; protein homodimerization activity; calcium-dependent phospholipid binding; phospholipase A2 inhibitor activity; calcium ion binding; calcium-dependent protein binding; helicase activity; protein binding; single-stranded RNA binding; double-stranded DNA-dependent ATPase activity; phospholipid binding; structural molecule activity; single-stranded DNA binding; receptor bindingBiological Process: prostate gland development; response to drug; response to peptide hormone stimulus; positive regulation of neutrophil apoptosis; gliogenesis; positive regulation of vesicle fusion; alpha-beta T cell differentiation; arachidonic acid secretion; neutrophil homeostasis; signal transduction; endocrine pancreas development; response to estradiol stimulus; DNA duplex unwinding; regulation of cell proliferation; keratinocyte differentiation; cell surface receptor linked signal transduction; insulin secretion; DNA strand renaturation; peptide cross-linking; cell motility; inflammatory response; response to X-ray; positive regulation of prostaglandin biosynthetic process; negative regulation of apoptosis
UniProt Protein Details:
NCBI Summary:
This gene encodes a membrane-localized protein that binds phospholipids. This protein inhibits phospholipase A2 and has anti-inflammatory activity. Loss of function or expression of this gene has been detected in multiple tumors. [provided by RefSeq, Dec 2014]
Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
Incubator
Deionized or distilled water
Absorbent paper
Buffer resevoir
*Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
Allow all reagents to reach room temperature (Please do not dissolve the reagents at 37°C directly). All the reagents should be mixed thoroughly by gently swirling before pipetting. Avoid foaming. Keep appropriate numbers of strips for 1 experiment and remove extra strips from microtiter plate. Removed strips should be resealed and stored at -20°C until the kits expiry date. Prepare all reagents, working standards and samples as directed in the previous sections. Please predict the concentration before assaying. If values for these are not within the range of the standard curve, users must determine the optimal sample dilutions for their experiments. We recommend running all samples in duplicate.
Step
1.
Add Sample: Add 100µL of Standard, Blank, or Sample per well. The blank well is added with Sample diluent. Solutions are added to the bottom of micro ELISA plate well, avoid inside wall touching and foaming as possible. Mix it gently. Cover the plate with sealer we provided. Incubate for 120 minutes at 37°C.
2.
Remove the liquid from each well, don't wash. Add 100µL of Detection Reagent A working solution to each well. Cover with the Plate sealer. Gently tap the plate to ensure thorough mixing. Incubate for 1 hour at 37°C. Note: if Detection Reagent A appears cloudy warm to room temperature until solution is uniform.
3.
Aspirate each well and wash, repeating the process three times. Wash by filling each well with Wash Buffer (approximately 400µL) (a squirt bottle, multi-channel pipette,manifold dispenser or automated washer are needed). Complete removal of liquid at each step is essential. After the last wash, completely remove remaining Wash Buffer by aspirating or decanting. Invert the plate and pat it against thick clean absorbent paper.
4.
Add 100µL of Detection Reagent B working solution to each well. Cover with the Plate sealer. Incubate for 60 minutes at 37°C.
5.
Repeat the wash process for five times as conducted in step 3.
6.
Add 90µL of Substrate Solution to each well. Cover with a new Plate sealer and incubate for 10-20 minutes at 37°C. Protect the plate from light. The reaction time can be shortened or extended according to the actual color change, but this should not exceed more than 30 minutes. When apparent gradient appears in standard wells, user should terminatethe reaction.
7.
Add 50µL of Stop Solution to each well. If color change does not appear uniform, gently tap the plate to ensure thorough mixing.
8.
Determine the optical density (OD value) of each well at once, using a micro-plate reader set to 450 nm. User should open the micro-plate reader in advance, preheat the instrument, and set the testing parameters.
9.
After experiment, store all reagents according to the specified storage temperature respectively until their expiry.
When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.
Sample Type
Protocol
Serum
If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.
Plasma
Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit.
Urine & Cerebrospinal Fluid
Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid.
Cell culture supernatant
Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately.
Cell lysates
Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
Tissue homogenates
The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C.
Tissue lysates
Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
Breast Milk
Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles.
