OPSS-PEG-NHS (molecular weight 5000) was purchased from Nanocs (New York, NY, USA). Trastuzumab (Herceptin) was a generous gift from Dr Virginia Borges (UC Denver, Denver, CO, USA). LLO from Escherichia coli transfected with the LLO-pEt29-DP-E3570 plasmid, kindly provided by Dr Dan Portnoy (UC Berkeley, Berkeley, CA, USA), was purified by the method described previously13 (link), 14 (link) and stored in storage buffer (50 mM phosphate buffer, pH 6.0, 1 M NaCl and 1 mM EDTA) without dithiothreitol to preserve its activity. Polylysine hydrobromide (molecular weight 37 000; degree of polymerization: 177) and 2-iminothiolane-HCl (Traut's reagent) were purchased from Sigma Life Science (St Louis, MO, USA). CL-4B Sepharose used for the purification of the one-component complexes was purchased from Amersham Biosciences (Uppsala, Sweden). All other reagents, unless otherwise specified, were purchased from Thermo Fisher Scientific (Pittsburgh, PA, USA).
2 iminothiolane hcl
Manufactured by Merck Group
Sourced in United States
2-iminothiolane-HCl is a chemical compound used in the laboratory for protein modification and cross-linking reactions. It is a heterobifunctional reagent that contains an amine-reactive group and a sulfhydryl-reactive group, allowing it to form covalent bonds between proteins or other biomolecules.
Automatically generated - may contain errors
Lab products found in correlation
Polylysine hydrobromide, by Merck Group (2 mentions)
Sepharose cl 4b, by Cytiva (2 mentions)
Traut s reagent, by Merck Group (1 mentions)
Fluorescamine, by Merck Group (1 mentions)
Pen strept, by Thermo Fisher Scientific (1 mentions)
Anti pdgfrα, by BioLegend (1 mentions)
Did solid, by Thermo Fisher Scientific (1 mentions)
Prestoblue cell viability reagent, by Thermo Fisher Scientific (1 mentions)
Propidium iodide, by Merck Group (1 mentions)
Siteclick antibody azido modification kit, by Thermo Fisher Scientific (1 mentions)
Labrafac, by Gattefosse (1 mentions)
4 protocols using 2 iminothiolane hcl
1
Trastuzumab-LLO Conjugation Protocol
2
Trastuzumab-Listeriolysin O Conjugate Synthesis
3
Targeted Delivery of Therapeutic Nanoparticles
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Labrafac® provided by Gattefosse SA (France) and Lipoïd® S100 by Lipoid Gmbh (Germany). Kolliphor HS15®, poly-d-Lysine hydrobromide (70000-150000 Da), fluorescamine, 2-Iminothiolane•HCl and propidium iodide were purchased from Sigma (United-States). 1,2-Distearoyl-sn-glycero-3-phosphorylethanolamine-PEG (2000) -Maleimide (DSPE-PEG-Maleimide) and NH 2 -PEG (5000) -Dibenzocyclooctyne (NH 2 -PEG-DBCO) were purchased from Nanocs (United-States). NH 2 -PEG (5000) -Maleimide was purchased from Creative PEGWorks (United-States). Anti-PDGFRα was purchased from BioLegend (United-States). Anti-rat-HRP, DMEM (41966-029), HEPES, Pen/Strept, DMEM 0.05% trypsin-EDTA (1x), Presto blue Cell Viability Reagent, DiD'solid, SiteClick™ Antibody Azido Modification Kit and Alexa-488-Phalloidine were purchased from Thermo Fisher Scientific (United-States). Plateletderived growth factor A (PDGFAA) and fibroblast growth factor 2 (FGF) were purchased from Peprotech (United Kingdom). rmPDGFRA/Fc chimera (soluble PDGFRα) has been purchased from R&Dsystems. Spectra-Por Float-a-lyzer G2 300 kDa dialysis membrane and Rotilabo® syringe sterile 0.22 μm filters were purchased from Carl Roth GmbH (Germany).
4
Synthesis of Partially Thiolated Chitosan
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Partially thiolated TMC (TMCSH) was prepared by reaction between the primary amino groups of purified polymer and 2-iminothiolane, as previously described (Scheme 1a) [14] . Briefly, TMC was dissolved in HEPES buffer (20 mM, pH 7.4) at 5 mg.ml-1. Then, 2-iminothiolane•HCl (Sigma-Aldrich, USA) was added in order to attain a theoretical modification of 10% of the polymer primary amines and the pH of the resulting solution adjusted to 8 with NaOH. The reaction mixture was saturated with argon and magnetically stirred at room temperature (RT) for 6 h. The resulting solution was dialyzed for 3 days against 5 mM HCl at 4 °C. Thereafter, the dialyzed solution was frozen at -80 °C and lyophilized for 3 days, yielding the TMCSH as a white powder, which was stored at -20 °C until further use.
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