The Rat Integrin Beta 1 (ITGB1) ELISA Kit is a powerful tool for the precise measurement of integrin beta 1 levels in rat serum, plasma, and cell culture supernatants. This kit offers exceptional sensitivity and specificity, ensuring accurate and consistent results for various research purposes.Integrin beta 1 is a key protein involved in cell adhesion and migration, playing a vital role in processes such as wound healing, inflammation, and cancer metastasis.
Understanding the expression levels of integrin beta 1 can provide valuable insights into these biological functions and help in the development of targeted therapies for related diseases.With its user-friendly protocol and high-quality components, the Rat Integrin Beta 1 (ITGB1) ELISA Kit is an indispensable tool for researchers studying integrin biology and its implications in various pathophysiological conditions.
Integrins alpha-1/beta-1, alpha-2/beta-1, alpha-10/beta-1 and alpha-11/beta-1 are receptors for collagen. Integrins alpha-1/beta-1 and alpha-2/beta-2 recognize the proline-hydroxylated sequence G-F-P-G-E-R in collagen. Integrins alpha-2/beta-1, alpha-3/beta-1, alpha-4/beta-1, alpha-5/beta-1, alpha-8/beta-1, alpha-10/beta-1, alpha-11/beta-1 and alpha-V/beta-1 are receptors for fibronectin. Alpha-4/beta-1 recognizes one or more domains within the alternatively spliced CS-1 and CS-5 regions of fibronectin. Integrin alpha-5/beta-1 is a receptor for fibrinogen. Integrin alpha-1/beta-1, alpha-2/beta-1, alpha-6/beta-1 and alpha-7/beta-1 are receptors for lamimin. Integrin alpha-4/beta-1 is a receptor for VCAM1 and recognizes the sequence Q-I-D-S in VCAM1. Integrin alpha-9/beta-1 is a receptor for VCAM1, cytotactin and osteopontin. It recognizes the sequence A-E-I-D-G-I-E-L in cytotactin. Integrin alpha-3/beta-1 is a receptor for epiligrin, thrombospondin and CSPG4. Integrin alpha-3/beta-1 provides a docking site for FAP (seprase) at invadopodia plasma membranes in a collagen-dependent manner and hence may participate in the adhesion, formation of invadopodia and matrix degradation processes, promoting cell invasion. Alpha-3/beta-1 may mediate with LGALS3 the stimulation by CSPG4 of endothelial cells migration. Integrin alpha-V/beta-1 is a receptor for vitronectin. Beta-1 integrins recognize the sequence R-G-D in a wide array of ligands. When associated with alpha-7/beta-1 integrin, regulates cell adhesion and laminin matrix deposition. Involved in promoting endothelial cell motility and angiogenesis. Involved in osteoblast compaction through the fibronectin fibrillogenesis cell-mediated matrix assembly process and the formation of mineralized bone nodules. May be involved in up-regulation of the activity of kinases such as PKC via binding to KRT1. Together with KRT1 and RACK1, serves as a platform for SRC activation or inactivation. Plays a mechanistic adhesive role during telophase, required for the successful completion of cytokinesis. ITGA4:ITGB1 binds to fractalkine (CX3CL1) and may act as its coreceptor in CX3CR1-dependent fractalkine signaling. ITGA4:ITGB1 and ITGA5:ITGB1 bind to PLA2G2A via a site (site 2) which is distinct from the classical ligand-binding site (site 1) and this induces integrin conformational changes and enhanced ligand binding to site 1. ITGA5:ITGB1 acts as a receptor for fibrillin-1 (FBN1) and mediates R-G-D-dependent cell adhesion to FBN1.
Uniprot:
P49134
Sample Type:
Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids
Specificity:
Natural and recombinant rat Integrin beta-1
Sub Unit:
Heterodimer of an alpha and a beta subunit. Beta-1 associates with either alpha-1, alpha-2, alpha-3, alpha-4, alpha-5, alpha-6, alpha-7, alpha-8, alpha-9, alpha-10, alpha-11 or alpha-V. Binds LGALS3BP and NMRK2, when associated with alpha-7, but not with alpha-5. Interacts with FLNA, FLNB, FLNC and RANBP9. Interacts with KRT1 in the presence of RACK1 and SRC. Interacts with JAML; integrin alpha-4/beta-1 may regulate leukocyte to endothelial cells adhesion by controlling JAML homodimerization. Interacts with RAB21. Interacts (via the cytoplasmic region) with RAB25 (via the hypervariable C-terminal region). Interacts with FGR and HCK. Interacts with MYO10. Interacts with DAB2. Interacts with ITGB1BP1 (via C-terminal region); the interaction is a prerequisite for focal adhesion disassembly. Interacts with FERMT2; the interaction is inhibited in presence of ITGB1BP1. Interacts with TLN1; the interaction is prevented by competitive binding of ITGB1BP1. Interacts with ACAP1; required for ITGB1 recycling. Interacts with ASAP3. Interacts with seprase FAP (seprase); the interaction occurs at the cell surface of invadopodia membrane in a collagen-dependent manner. Interacts with EMP2; the interaction may be direct or indirect and ITGB1 has a heterodimer form (By similarity). ITGA5:ITGB1 interacts with NOV. ITGA4:ITGB1 is found in a ternary complex with CX3CR1 and CX3CL1. ITGA5:ITGB1 interacts with FBN1.
Research Area:
Cancer
Subcellular Location:
Cell membrane Single-pass type I membrane protein Cell projection Invadopodium membrane Single-pass type I membrane protein Cell projection Ruffle membrane Single-pass type I membrane protein Recycling endosome Melanosome Cell projection Lamellipodium Cell projection Ruffle Cell junction Focal adhesion Cell surface Enriched preferentially at invadopodia, cell membrane protrusions that correspond to sites of cell invasion, in a collagen-dependent manner. Localized at plasma and ruffle membranes in a collagen-independent manner. Colocalizes with ITGB1BP1 and metastatic suppressor protein NME2 at the edge or peripheral ruffles and lamellipodia during the early stages of cell spreading on fibronectin or collagen. Translocates from peripheral focal adhesions to fibrillar adhesions in an ITGB1BP1-dependent manner.
Storage:
Please see kit components below for exact storage details
Note:
For research use only
UniProt Protein Function:
ITGB1: an integral membrane protein that heterodimerizes with an alpha-3 chain, forming a receptor for many extracellular-matrix proteins including fibronectin, laminin, collagen, epiligrin and thrombospondin. . Beta 1 integrins recognize the amino-acid motif RGD in a wide array of ligands. Five alternatively spliced variants with alternate carboxy termini have been described. Two alternatively spliced isoforms have been described. Isoform beta-1a is widely expressed; other isoforms are generally expressed with a more restricted distribution. Isoform beta-1b is expressed in skin, liver, skeletal muscle, cardiac muscle, placenta, umbilical vein endothelial cells, neuroblastoma cells, lymphoma cells, hepatoma cells and astrocytoma cells. Isoforms beta-1c and beta-1c-2 are expressed in muscle, kidney, liver, placenta, cervical epithelium, umbilical vein endothelial cells, fibroblast cells, embryonal kidney cells, platelets and several blood cell lines. Isoform beta-c-2, rather than isoform beta-1c, is selectively expressed in primary t-cells. Isoform beta-1c is expressed in nonproliferating and differentiated prostate gland epithelial cells. Isoform beta-1d is expressed specifically in striated muscle (skeletal and cardiac muscle).Protein type: Motility/polarity/chemotaxis; Cell adhesion; Receptor, misc.; Membrane protein, integral; Cell surfaceChromosomal Location of Human Ortholog: 10p11.2Cellular Component: focal adhesion; cell surface; dendritic spine; acrosome; lipid raft; ruffle; recycling endosome; hemidesmosome; membrane; lamellipodium; cytoplasm; plasma membrane; melanosome; basement membrane; integrin complex; neuromuscular junction; sarcolemma; receptor complex; external side of plasma membrane; cleavage furrow; filopodiumMolecular Function: protein domain specific binding; protease binding; metal ion binding; laminin binding; actin binding; alpha-actinin binding; protein kinase binding; peptide binding; viral receptor activity; integrin binding; protein binding; protein heterodimerization activity; fibronectin binding; cell adhesion molecule binding; glycoprotein bindingBiological Process: extracellular matrix organization and biogenesis; maternal process involved in pregnancy; regulation of cell cycle; axon extension; positive regulation of apoptosis; mesodermal cell differentiation; positive regulation of endocytosis; cardiac muscle cell differentiation; leukocyte adhesion; transforming growth factor beta receptor signaling pathway; germ cell migration; tissue homeostasis; response to drug; dendrite morphogenesis; protein transport within lipid bilayer; positive regulation of peptidyl-tyrosine phosphorylation; negative regulation of neuron differentiation; cell migration during sprouting angiogenesis; B cell differentiation; cell-cell adhesion mediated by integrin; response to activity; leukocyte migration; regulation of G-protein coupled receptor protein signaling pathway; G1/S transition of mitotic cell cycle; axon guidance; entry of virus into host cell; cell-matrix adhesion; cell fate specification; negative regulation of cell proliferation; positive regulation of MAPKKK cascade; positive regulation of cell proliferation; visual learning; negative regulation of cell projection organization and biogenesis; integrin-mediated signaling pathway; regulation of immune response; cell migration; in utero embryonic development; sarcomere organization; cell-substrate adhesion; formation of radial glial scaffolds; cellular calcium ion homeostasis; heterotypic cell-cell adhesion; negative regulation of Rho protein signal transduction; calcium-independent cell-matrix adhesion; stress fiber formation; cellular defense response; homophilic cell adhesion; blood coagulation; leukocyte tethering or rolling; positive regulation of cell migration
UniProt Protein Details:
NCBI Summary:
Integrins are heterodimeric proteins made up of alpha and beta subunits. At least 18 alpha and 8 beta subunits have been described in mammals. Integrin family members are membrane receptors involved in cell adhesion and recognition in a variety of processes including embryogenesis, hemostasis, tissue repair, immune response and metastatic diffusion of tumor cells. This gene encodes a beta subunit. Multiple alternatively spliced transcript variants which encode different protein isoforms have been found for this gene. [provided by RefSeq, Jul 2008]
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.
