Key resources table.
| Reagent type (species) or resource |
Designation | Source or reference |
Identifiers | Additional information |
|---|---|---|---|---|
| Recombinant DNA reagent | pMT-puro VWFA1-V5-His | In house | The insect cell expression vector to express VWF A1 domain can be obtained from Prof Crawley | |
| Recombinant DNA reagent | pMT-puro VWFA1*- V5-His | In house | The insect cell expression vector to express VWF A1* domain can be obtained from Prof Crawley | |
| Bacterial strain, strain background (E. coli) | TOP10 (DH10B background) |
Invitrogen | C4040-03 | Chemically competent cells |
| Antibody | anti-6xHis-HRP (rabbit polyclonal) | Abcam | RRID:AB_298652 | (WB: 1:10000) |
| Cell line (D. melanogaster) | S2 | S2 cells originally from ATCC transfected and selected in house | VWF A1-V5-His/S2 and VWF A1*-V5-His/S2 | Cell line maintained in Crawley lab; stably transfected with human VWF A1 domain-V5-Hisor VWF A1*-V5-His |
| Cell line (Homo-sapiens) | Human embryonic Kidney HEK293T | ATCC | RRID:CVCL_0063 | |
| Antibody | anti-VWF (rabbit polyclonal) | Agilent | RRID:AB_2315602 | (WB 1:1000) |
| Biological sample (Homo sapiens) | αIIbβ3 | Enzyme Research Laboratories | GP2b3a | αIIbβ3 isolated from human platelets |
| Antibody | anti-PECAM-1 (mouse monoclonal) | BioLegend | RRID:AB_314328 | (flow assays: 0.25 mg/ml IF:50 μg/ml) |
| Antibody | anti-beta3 (LIBS2) (mouse monoclonal) | Millipore | RRID:AB_10806476 | (flow assays 0.25 mg/ml) |
| Antibody | anti-CD16 (mouse monoclonal) | eBiosciences | RRID:AB_467129 | (flow assays 0.25 mg/ml) |
| Chemical compound, drug | eptifibatide | Sigma | SML1042-10MG | αIIbβ3 blocking/inhibition 2.4 μM |
| Chemical compound, drug | GR144053 | Tocris | 1263 | αIIbβ3 blocking/inhibition 2 μM |
| Antibody | anti-P-selectin (mouse monoclonal) | BD Biosciences | RRID:AB_395908 | Clone Ak4 (blocking P-selectin: 50 μg/ml) |
| Antibody | anti-SLC44A2 #1 (rabbit polyclonal) | Abcam | RRID:AB_2827953 | (SLC44A2 blocking 0–20 μg/ml) |
| Antibody | anti-SLC44A2 #2 (rabbit polyclonal) | LS Bio | RRID:AB_2827954 | (SLC44A2 blocking 0–20 μg/ml) |
| Antibody | anti-CD14-APC (mouse monoclonal) | BioLegend | RRID:AB_314190 | (IF 1:20) |
| Antibody | anti-CD3-APC (mouse monoclonal) | BioLegend | RRID:AB_314066 | (IF 1:20) |
| Antibody | anti-CD19-APC (mouse monoclonal) | BioLegend | RRID:AB_314242 | (IF 1:20) |
| Antibody | anti-CD16-APC (mouse monoclonal) | eBiosciences | RRID:AB_2016663 | (IF 1:20) |
| Chemical compound, drug | TMB-8 | Sigma | T111 | PKC inhibition: 20 μM |
| Chemical compound, drug | DPI | Sigma | 300260 | NADPH oxidase inhibition: 30 μM |
| Recombinant DNA reagent | pCMV6 Entry/SLC44A2-tGFP (plasmid with cDNA) | Origene | RG207181 | Expression vector of human SLC44A2 with C-terminal turbo GFP fusion |
| Recombinant DNA reagent | pCMV6 Entry/SLC44A2(R154Q)-tGFP (plasmid with cDNA) | In house | As above but containing SLC44A2 (R154Q) variant. The expression vector pCMV6 Entry/SLC44A2(R154Q)-tGFP can be obtained from Prof Crawley | |
| Sequence-based reagent | R154Q top primer | Invitrogen | 5’-GTG GCT GAG GTG CTT CAA GAT GGT GAC TGC CCT-3’ | |
| Sequence-based reagent | R154Q bot primer | Invitrogen | 5’-AGG GCA GTC ACC ATC TTG AAG CAC CTC AGC CAC-3’ | |
| Sequence-based reagent | SLC44A2 top sequencing primer | Invitrogen | 5’-ACC TCA CGT ACC TGA ATG-3’ | |
| Sequence-based reagent | SLC44A2 bot sequencing primer | Invitrogen | 5’-AGC CAT GCC CAT CCT CAT AG-3’ | |
| Antibody | anti-tGFP (mouse monoclonal) | Origene | RRID:AB_2622256 | (WB:0.3 μg/ml) |
| Antibody | anti-Cit H3 (rabbit polyclonal) | Abcam | RRID:AB_304752 | (IF:10 μg/ml) |
| Software | GraphPad Prism | GraphPad Prism | RRID:SCR_002798 | |
| Software | SlideBook 6.0 | 3i | RRID:SCR_014300 |