Specific rabbit antibodies were obtained by immunizing rabbits with 200 µg aliquots of recombinant ICAM-1 (R&D Systems, Minneapolis, MN, USA) and purified recombinant VP2, VP3, and VP4 that had been purified as described [15 (link)]. The first immunizations were conducted with a mix of the respective protein and Freund’s Complete Adjuvant (CFA) (Charles River, Chatillon sur Chalaronnne, France) and the 2nd to 4th immunization with a mix of each protein and Freund’s Incomplete Adjuvant (IFA) (Charles River, Chatillon sur Chalaronnne, France). Rabbit anti-VP1 antibodies were as previously described [27 (link)]. Commercial rabbit anti-ICAM-1 (Sino Biological, Beijing, China) and goat anti-LDLR antibodies (R&D Systems, Minneapolis, MN, USA) used in certain experiments were purchased. Normal rabbit antibodies used for control experiments were obtained from Genscript (Leiden, Netherlands). RV-strain-specific guinea pig antibodies were purchased from ATCC.
Recombinant icam 1
Manufactured by R&D Systems
Sourced in United States, France
Recombinant ICAM-1 is a protein produced in a laboratory setting. It is a type of cell adhesion molecule that plays a role in cell-to-cell interactions. The core function of Recombinant ICAM-1 is to facilitate the binding and adhesion of cells.
Automatically generated - may contain errors
Lab products found in correlation
4 protocols using recombinant icam 1
1
Producing Strain-Specific Antibodies for Immunological Research
2
Immobilizing Protein Coatings for AFM
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A 20 µL aliquot of recombinant ICAM-1 (RD Systems, Lille, France) (25 µg/ml) in 0.1 M NaHCO3 (pH 8.6) was adsorbed overnight at 4°C at the centre of the coverslip. Unbound proteins were removed by washing with PBS and 2 ml of complete RPMI 1640 medium were then added to the ICAM-1 coated dish before the AFM experiments.
For the BSA-coating protocol, a µL aliquot of BSA at 100 µg/ml in PBS was adsorbed 30 minutes at 37°C at the centre of the Petri dish. Unbound proteins were removed by washing with PBS and 2 ml of the complete RPMI medium were then added to the BSA coated dish before the AFM experiments.
For the BSA-coating protocol, a µL aliquot of BSA at 100 µg/ml in PBS was adsorbed 30 minutes at 37°C at the centre of the Petri dish. Unbound proteins were removed by washing with PBS and 2 ml of the complete RPMI medium were then added to the BSA coated dish before the AFM experiments.
3
Recombinant ICAM1 and Phl p Allergen ELISA
4
Neutrophil Adhesion Assay using ICAM-1
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Individual wells of Immulon2HB flat bottom 96-well plates were coated with 10 μg/mL Recombinant ICAM-1 (R&D Systems) or 5% FBS in PBS (for controls). Isolated human neutrophils at 1 × 107/mL in HBSS were labeled with calcein AM (Corning) at a concentration of 2 μg/mL and primed with human GM-CSF (EMD Millipore) at 5 ng/mL for 30 minutes, protected from light. Cells were then centrifuged at 1000 rpm for 8 minutes and then resuspended in HBSS++ +2% FBS buffer at 7.0 × 105 cells/mL. Appropriate blocking antibodies and/or peptides were added to cells and incubated for 30 minutes at 37°C. The ICAM-1 coated plate was washed once with 1X PBS before cells were added to individual wells. Cells were added to wells and allowed to settle for 10 minutes at 37°C. 0.5 mM Mn2+ was added to individual wells. Plates were incubated for 10 minutes prior to initial fluorescence reading. Cells were gently dumped and washed with PBS, reading fluorescence (485 nm excitation, 535 nm emission) at each wash step. Fluorescence washing was divided by initial fluorescence and multiplied by 100 to calculate percent adhesion. The first wash that demonstrated less than 10% adhesion in the non-stimulated cells (plated on 5% FBS coating) was considered the result. Treatment groups were tested in triplicate.
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