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Vitronectin

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Vitronectin is a glycoprotein that plays a role in cell adhesion, migration, and tissue remodeling. It serves as a substrate for cell attachment and can modulate the activity of various cell surface receptors. Vitronectin is a component of the extracellular matrix and is involved in a variety of biological processes.

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Lab products found in correlation

Fibronectin, by Roche (4 mentions) Cm7 chip, by GE Healthcare (4 mentions) Recombinant human integrin αvβ3, by R&D Systems (4 mentions) Fibronectin, by R&D Systems (3 mentions) Acetate buffer ph5, by GE Healthcare (3 mentions) Diff quick solutions, by Thermo Fisher Scientific (2 mentions) Fibronectin, by BD (2 mentions) Axioskop 2 plus microscope, by Zeiss (1 mentions) Type 1 collagen, by BD (1 mentions) Collagen type 2, by BD (1 mentions) Spectramax gemini xs, by Molecular Devices (1 mentions) Gelatin, by BD (1 mentions) Pten activity elisa, by Echelon Biosciences (1 mentions) Rmfg e8, by R&D Systems (1 mentions) Matrigel, by BD (1 mentions) Collagen 4, by R&D Systems (1 mentions) Collagen 1, by Thermo Fisher Scientific (1 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions) L glutamine, by Thermo Fisher Scientific (1 mentions) 4 6 diamidino 2 phenylindole dapi nuclear stain, by Vector Laboratories (1 mentions)

Spelling variants of this product

Human vitronectin (3 citations)

17 protocols using vitronectin

1

Vitronectin-Mediated Cell Adhesion Assay

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OC precursors were seeded in 48-wells plates coated with 10 μg/ml vitronectin (R&D Systems, Minneapolis, MN) for 10 and 60 min at 37°C. Unattached cells were rinsed with PBS, stained with Diff-Quick solutions (Thermo, Rockford, IL), observed under the Axioskop2 Plus microscope (Carl Zeiss, Oberkochen, Germany), and then the number of adhered cells was counted.
Jang J.S., Kang I.S., Cha Y.N., Lee Z.H., Dinauer M.C., Kim Y.J, & Kim C. (2019). Vav1 inhibits RANKL-induced osteoclast differentiation and bone resorption. BMB Reports, 52(11), 659-664.
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2

Extracellular Matrix Adhesion Assay

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Wells of 96 well plates were coated with 1% gelatin, 10 μg/mL collagen type I, 10μg/mL collagen type II (BD Biosciences), 10 μg/mL fibronectin (BD Biosciences), 10 μg/ mL fibrinogen (gift from Dr. Edward Plow), or 10 μg/mL vitronectin (R&D Systems) at 37 °C overnight and then blocked with 0.1% BSA for 1 hour prior to plating. We used BSA coated wells as a control for weak substrate attachment. Before plating cells were incubated in a calcein-AM green solution (Invitrogen) for 30 minutes at 37°C. Cells were allowed to attach for one hour. Calcein fluorescence was read at 485 nm excitation and 538 nm emission (SpectraMAX Gemini XS, Molecular Devices). Values were normalized to the BSA control. Experiments were repeated 3 times.
Kerr B.A., Shi L., Jinnah A.H., Harris K.S., Willey J.S., Lennon D.P., Caplan A.I, & Byzova T.V. (2021). Kindlin-3 Mutation in Mesenchymal Stem Cells Results in Enhanced Chondrogenesis. Experimental cell research, 399(2), 112456.
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3

Adhesion Assay of Breast Cancer Cells

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The adhesion assay was performed using 96-well microplates coated with vitronectin (R&D Systems, Wiesbaden, Germany), gelatin, collagen or fibronectin (all from BD, Heidelberg, Germany). The microplates were rehydrated with 200 μl PBS/well for 30 min at room temperature prior to use and the PBS removed before adding the cells. MDA-231 breast cancer cells were treated with different inhibitors of the mevalonate pathway for 24 h as indicated. Cells were then stained with the fluorescent dye DilC12(3) (from BD Biosciences, Heidelberg, Germany) for 1 h and washed twice with PBS. DilC12(3) is a fluorescent tracer specifically designed to label viable cells for tumor cell invasion or migration assays. After carefully harvesting the cells with 0.0015 M EDTA/PBS, they were washed again with PBS, counted and reconstituted in DMEM. Cells (125,000/well) were then given on 96-well microplates coated with different surfaces and incubated at 37 °C for 1 h in a CO2 incubator allowing them to adhere. Afterwards the plates were washed gently 3 times with PBS to remove non adherent cells and 100 μl of DMEM added into the wells. The adherent cells were then quantified by their relative fluorescence signal with Fluostar Omega plate reader at 544/590 nm.
Wilke M., Göbel A., Rauner M., Benad-Mehner P., Schütze N., Füssel S., Hadji P., Hofbauer L.C, & Rachner T.D. (2014). Zoledronic acid and atorvastatin inhibit αvβ3-mediated adhesion of breast cancer cells. Journal of Bone Oncology, 3(1), 10-17.
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4

PTEN Activity Modulation by ECM Proteins

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We isolated antral lysates or human gastric smooth muscle cell lysates and measured conversion of PI(3,4,5) P3 to PI(4,5)P2 (PTEN activity ELISA, Echelon) after incubation with recombinant proteins (rMfge8, RGE, Fibronectin, or Vitronectin, R&D Systems, Inc. 10 μg/ml) or blocking antibodies against α8, β3, and β5 (10 g/ml). In this competitive ELISA, we incubated lysates on a PI(4,5)P2 coated microplate and then added a PI(4,5)P2 detector protein. PI(4,5)P2 produced by PTEN in lysate binds to the detector protein and thus prevents it from binding immobilized PI(4,5)P2 on the plate. We then used a peroxidase-linked secondary to measure PI(4,5)P2 detector protein binding to the plate in a colorimetric assay where the colorimetric signal is inversely proportional to the amount of PI(4,5)P2 produced by PTEN.
Khalifeh-Soltani A., Ha A., Podolsky M.J., McCarthy D.A., McKleroy W., Azary S., Sakuma S., Tharp K.M., Wu N., Yokosaki Y., Hart D., Stahl A, & Atabai K. (2016). α8β1 integrin regulates nutrient absorption through an Mfge8-PTEN dependent mechanism. eLife, 5, e13063.
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5

Quantifying Cell Adhesion Mediated by uPAR

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96-well plates were coated with 5 ng•µl−1 vitronectin (R&D, Minneapolis, MN, USA) at 4 °C overnight, and then blocked with 2 % bovine serum albumin (BSA) in PBS for 1 h. After starving with serum-free medium for 4 h, 293-uPAR and HEK293 cells (2.5×104 cells mL−1) were suspended in 100 µL of DMEM containing 0.1 % FBS with indicated concentrations of IPR1110, or DMSO control in the presence or absence of 500pM uPA-ATF at 37 °C for 90 min. Medium was then carefully suctioned out from each well. Each well was washed three times with PBS. After washing, the adherent cells were fixed, stained with crystal violet, and quantified by measuring the absorbance at 540 nm.
Liu D., Zhou D., Wang B., Knabe W.E, & Meroueh S.O. (2015). A New Class of Orthosteric uPAR•uPA Small-Molecule Antagonists Are Allosteric Inhibitors of the uPAR•Vitronectin Interaction. ACS chemical biology, 10(6), 1521-1534.
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6

Cell Adhesion Assay on ECM Proteins

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96-well plates were coated overnight at 4°C with vitronectin (1 μg/cm2; R&D Systems), fibronectin (10 μg/cm2; R&D Systems), Matrigel (5 μg/cm2; BD Biosicences) or collagen IV (10 μg/cm2; R&D Systems) and then blocked for 1 h with 0.5% bovine serum albumin. Cells were seeded at a density of 4 × 104 cells/well and incubated at 37°C for 1.5 h. Non-adherent cells were removed by washing with PBS, and adherent cells were fixed with cold methanol and stained with 0.1% crystal violet for 25 min at room temperature. After removing the crystal violet solution, the stained cells were washed with water and 10% acetic acid was added to dissolve the crystal violet. Absorbances were measured at 590 nm using a microplate spectrophotometer.
Xiong S., Klausen C., Cheng J.C., Zhu H, & Leung P.C. (2015). Activin B induces human endometrial cancer cell adhesion, migration and invasion by up-regulating integrin β3 via SMAD2/3 signaling. Oncotarget, 6(31), 31659-31673.
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7

Cell Adhesion Dynamics on Coated Plates

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Ratio of adherent versus floating cells at various times points after seeding was measured for SB.mhdgc-1 and SB.msgc-1 cells on regular 24-well tissues culture plates, and on 24-well plates coated with collagen I, fibronection, laminin I (all ThermoFisher Scientific, Waltham, MA), and 96-well plates coated with vitronectin (R&D Systems, Minneapolis, MN). Cells were seeded in six replicates at 1.0 × 105 cells per well in 24-well plates and 2 × 104 cells per well in 96-well plates and two independent cell counts in Nexcelom Auto T4 Hemacytometer cell chambers (Nexcelom, Lawrence, MA) were obtained. Time course ratios of adherent versus floating cells were graphed using GraphPad Prism version 7 (La Jolla, CA).
Chen I., Mathews-Greiner L., Li D., Abisoye-Ogunniyan A., Ray S., Bian Y., Shukla V., Zhang X., Guha R., Thomas C., Gryder B., Zacharia A., Beane J.D., Ravichandran S., Ferrer M, & Rudloff U. (2017). Transcriptomic profiling and quantitative high-throughput (qHTS) drug screening of CDH1 deficient hereditary diffuse gastric cancer (HDGC) cells identify treatment leads for familial gastric cancer. Journal of Translational Medicine, 15, 92.
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8

Integrin αVβ3 Binding Assay

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Example 13

Recombinant human integrin αVβ3 (R&D Systems, Minneapolis Minn.) was diluted to 40 ug/mL in Acetate buffer pH5.0 (GE Healthcare, Piscataway N.J.), and. then immobilized on a CM7 chip (GE Healthcare) using standard amine coupling techniques. 500 nM fibronectin (Roche Diagnostics, Indianapolis, Ind.) and vitronectin (R&D Systems) and 5 μM of either non-binding control 10Fn3 molecule (consisting of SEQ ID NO: 6 with an additional MG at the N-terminus and with a single amino acid substitution that changes RGD to RGE) or targeted 10Fn3 molecules (having a mutated FG loop that does not contain an RGD motif) were flowed over the top of the immobilized integrin. Binding RU was collected at the end of the sample injection. The results indicate that the lack of RGD in the FG loop results in abolishing binding of 10Fn3 molecules to fibronectin and vitronectin.

US10604556B2. Fibronectin binding domains with reduced immunogenicity (2020-03-31). BRISTOL-MYERS SQUIBB COMPANY [US]. Inventors: Jonathan Davis [US], Dasa Lipovsek [US], Ray Camphausen [US].
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9

Integrin αVβ3 Binding Inhibition

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Example 13

Recombinant human integrin αVβ3 (R&D Systems, Minneapolis Minn.) was diluted to 40 ug/mL in Acetate buffer pH5.0 (GE Healthcare, Piscataway N.J.), and. then immobilized on a CM7 chip (GE Healthcare) using standard amine coupling techniques. 500 nM fibronectin (Roche Diagnostics, Indianapolis, Ind.) and vitronectin (R&D Systems) and 5 μM of either non-binding control 10Fn3 molecule (consisting of SEQ ID NO: 6 with an additional MG at the N-terminus and with a single amino acid substitution that changes RGD to RGE) or targeted 10Fn3 molecules (having a mutated FG loop that does not contain an RGD motif) were flowed over the top of the immobilized integrin. Binding RU was collected at the end of the sample injection. The results indicate that the lack of RGD in the FG loop results in abolishing binding of 10Fn3 molecules to fibronectin and vitronectin.

US10464995B2. Fibronectin binding domains with reduced immunogenicity (2019-11-05). BRISTOL-MYERS SQUIBB COMPANY [US]. Inventors: Jonathan Davis [US], Dasa Lipovsek [US], Ray Camphausen [US].
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10

Integrin αVβ3 Binding Characterization

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Example 13

Recombinant human integrin αVβ3 (R&D Systems, Minneapolis Minn.) was diluted to 40 ug/mL in Acetate buffer pH5.0 (GE Healthcare, Piscataway N.J.), and. then immobilized on a CM7 chip (GE Healthcare) using standard amine coupling techniques. 500 nM fibronectin (Roche Diagnostics, Indianapolis, Ind.) and vitronectin (R&D Systems) and 5 μM of either non-binding control 10Fn3 molecule (consisting of SEQ ID NO: 6 with an additional MG at the N-terminus and with a single amino acid substitution that changes RGD to RGE) or targeted 10Fn3 molecules (having a mutated FG loop that does not contain an RGD motif) were flowed over the top of the immobilized integrin. Binding RU was collected at the end of the sample injection. The results indicate that the lack of RGD in the FG loop results in abolishing binding of 10Fn3 molecules to fibronectin and vitronectin.

US11447538B2. Fibronectin based scaffold proteins (2022-09-20). BRISTOL-MYERS SQUIBB COMPANY [US]. Inventors: Dasa Lipovsek [US], Jonathan H. Davis [US].
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