Isobutylcyanoacrylate (IBCA) was purchased from Orapi (Saint-Vulbas, France). Dextran 17.7 kDa and cerium (IV) ammonium nitrate were provided by Fluka (Saint-Quentin-Fallavier, France). Dextran 66.7 kDa, Tricine, Tris base (Sigma 7-9 ® ), Sodium chloride, EDTA sodium salt and Bromophenol blue were purchased from Sigma (Saint-Quentin-Fallavier, France). Nitric acid was supplied by Prolabo (Paris, France). Human serum was prepared from plasma provided by Etablissement Français du Sang (EFS) (Rungis, France). Polyclonal anti-human C3 antibody raised in goat was purchased from Fitzgerald antibodies (Acton, USA). Coomassie brilliant blue R-250, calcium lactate and glacial acetic acid were supplied by Thermo Fisher Scientific (Villebon-sur-Yvette, France). Gel-Fix™ for agarose (265x150 mm) was purchased from Serva Electrophoresis (Heidelberg, Germany). Nafamostat mesylate (NM) was obtained from Abcam (Cambridge, UK).
Nafamostat mesylate
Manufactured by Abcam
Sourced in United Kingdom
Nafamostat mesylate (NM) is a synthetic serine protease inhibitor. It is commonly used in research applications to inhibit trypsin-like proteases.
Automatically generated - may contain errors
Lab products found in correlation
Glacial acetic acid, by Thermo Fisher Scientific (2 mentions)
Coomassie brilliant blue r 250, by Thermo Fisher Scientific (2 mentions)
Calcium lactate, by Thermo Fisher Scientific (2 mentions)
Bromophenol blue, by Merck Group (2 mentions)
Dextran 66.7 kda, by Merck Group (2 mentions)
Tricine, by Merck Group (2 mentions)
Edta sodium salt, by Merck Group (1 mentions)
Tris base, by Merck Group (1 mentions)
Batimastat, by Merck Group (1 mentions)
Varespladib, by Merck Group (1 mentions)
Dimercaprol, by Merck Group (1 mentions)
Marimastat, by Merck Group (1 mentions)
4 protocols using nafamostat mesylate
1
Nanoparticle Synthesis and Characterization
2
Isobutylcyanoacrylate Biomedical Protocol
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Isobutylcyanoacrylate (IBCA) was synthesized and provided by Orapi (Saint-Vulbas, France). Chemicals purchased from Sigma (Saint-Quentin-Fallavier, France) were tricine, Tris base (Sigma 7-9 ® ), sodium chloride, EDTA sodium salt, dextran 66.7 kDa, and bromophenol blue. Dextran 17.7, cerium (IV) ammonium nitrate, nitric acid, and chloride acid were supplied by Fluka (Saint-Quentin-Fallavier, France). Nitric acid was purchased from Prolabo (Paris, France). Calcium lactate, glacial acetic acid, coomassie brilliant blue R-250 were supplied by Thermo Fisher Scientific (Villebon-sur-Yvette, France). Gel-Fix™ for agarose (265x150 mm) was obtained from Serva Electrophoresis (Heidelberg, Germany). Nafamostat mesylate (NM) was obtained from Abcam (Cambridge, UK). All chemicals were of reagent grade and used as purchased.
Polyclonal anti-human C3 antibody raised in goat was purchased from Fitzgerald antibodies (Acton, USA). Serum was prepared from human plasma obtained from healthy donors from Etablissement Français du Sang (EFS) (Rungis, France). Heat-activated gamma globulin (HAGG) and Cobra Venom Factor (CVF) were purchased from TECO Medical (Rambouillet, France) to be used as positive control for the activation of the complement system.
Polyclonal anti-human C3 antibody raised in goat was purchased from Fitzgerald antibodies (Acton, USA). Serum was prepared from human plasma obtained from healthy donors from Etablissement Français du Sang (EFS) (Rungis, France). Heat-activated gamma globulin (HAGG) and Cobra Venom Factor (CVF) were purchased from TECO Medical (Rambouillet, France) to be used as positive control for the activation of the complement system.
3
Preparation of Small Molecule Inhibitors
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Dimercaprol (2,3-dimercapto-1-propanol ≥98% iodometric, Cat no: 64046-10 ml), marimastat (≥98% HPLC M2699-5MG), batimastat (SML0041-5MG) and varespladib (≥98% HPLC SML1100-5MG) were purchased from Sigma-Aldrich. Nafamostat mesylate (ab141432 10 mg) was purchased from Abcam and DMPS (2,3-dimercapto-1-propanesulfonic acid sodium salt monohydrate, 95%, Cat no: H56578) from Alfa Aesar. Working stocks (tenfold dilutions from 2 mM to 2 µM) were made using deionized water, with the exception of varespladib and batimastat, for which we used DMSO due to water insolubility.
4
Chlamydia trachomatis Complement Activation
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Reactions were carried out at 37°C with a reaction volume of 30 μL in a 1.5-mL Eppendorf tube containing 400 nM purified human C5 (Complement Technologies, Tyler, TX, USA) mixed with 25 μL C. trachomatis-HeLa229 cell lysates (C. trachomatis infection group), mock-infected HeLa229 cell lysates (mock infection group), PBS (C5 control group) or EBs. In the inhibition assay, C. trachomatis-HeLa229 cell lysates were pre-mixed with 400 μM nafamostat mesylate (Abcam, Cambridge, MA, USA) for 10 min, or with 500 μM lactacystin (Glpbio, Montclair, CA, USA) for 30 min respectively, both at 37°C. Reactions were terminated at various time points as indicated for the individual experiments by immediately placed on ice. Cleavage products were subjected to Western blot analysis, enzyme-linked immunosorbent assay (ELISA), and C-terminal sequencing based on liquid chromatography-tandem mass spectrometry (LC-MS/MS).
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