Table 1.
Antibodies Used for Immunohistochemistry
| Antibody, Clone or Catalog No. | Abbreviation Used in Study | Host Isotype; Subclass | Purification | Cellular Localization | Pretreatment | Protein Block (Dako) | Dilution | Secondary Antibody | Source |
|---|---|---|---|---|---|---|---|---|---|
| Vimentin, 3B4 | Vim |
Mouse IgG2a |
Purified from cell culture supernatant | Cytoskeleton (intermediate filament) | Tris‐EDTA (pH 9.2) | No | 1:2000 |
|
Dako |
| Fibroblast‐specific protein‐1/S100A4, D9F9D | FSP‐1 | Rabbit IgG | Not specified | Nucleus, cytoplasm, and extracellular space | Tris‐EDTA (pH 9.2) | No | 1:6000 | DAKO EnVision+ System, anti‐rabbit | Cell Signaling Technology |
| CD90/thymocyte differentiation antigen 1, ERP3133 | Thy‐1 | Rabbit IgG | Purified from tissue culture supernatant | Cell membrane | Tris‐EDTA (pH 9.2) | No | 1:200 | DAKO EnVision+ System, anti‐rabbit | Abcam |
| Fibroblast activation protein, AF3715 | FAP | Sheep IgG | Antigen affinity purified | Cell membrane | Tris‐EDTA (pH 9.2) | No | 1:40 | Donkey anti‐sheep IgG‐HRP, A16047 (ThermoFisher) | R&D Systems |
| α‐Smooth muscle actin, 1A4 | αSMA | Mouse IgG2a | Purified from cell culture supernatant | Cytoskeleton (actin filaments) | Tris‐EDTA (pH 9.2) | No | 1:4000 |
|
Dako |
| Nonmuscle myosin heavy chain, NBP2‐38566 | Smemb | Rabbit IgG | Immunogen affinity purified | Cytoskeleton (myosin binding) | Tris‐EDTA (pH 9.2) | No | 1:600 | DAKO EnVision+ System, anti‐rabbit | Novusbio |
| Smooth muscle myosin heavy chain I (SM1), 3F8 | SM‐MHC (SM1) |
Mouse IgG1 |
Ascites | Cytoskeleton (myosin) | Tris‐EDTA (pH 9.2) | No | 1:5000 | DAKO EnVision+ System, anti‐mouse | Abcam |
| Smooth muscle myosin heavy chain 11 (SM2), ab53219 | SM‐MHC (SM2) | Rabbit IgG | Ion exchange chromatography | Cytoskeleton (myosin) | Tris‐EDTA (pH 9.2) | No | 1:500 | DAKO EnVision+ System, anti‐rabbit | Abcam |
| Calponin, hCO | Calp | Mouse IgG1 | Ascites | Cytoskeleton (thin filament‐associated protein) | Tris‐EDTA (pH 9.2) | Yes | 1:3 000 000 |
|
Merck |
| Caldesmon, h‐CALD | h‐Caldes |
Mouse IgG1 |
Protein A or G | Cytoskeleton (microfilaments) and ECM | Citrate (pH 6) | No | 1:2000 | DAKO EnVision+ System, anti‐mouse | Novusbio |
| Desmin, AF3844 | Des | Goat IgG | Antigen affinity purified | Cytoskeleton (intermediate filament) | Tris‐EDTA (pH 9.2) | No | 1:50 | Donkey anti‐goat IgG‐HRP, ab97120 (Abcam) | R&D Systems |
| Smooth muscle protein 22α/transgelin, 8C8 | SM22α | Mouse IgG1 | Antigen affinity purified | Cytoskeleton (actin binding) | No retrieval | No | 1:25000 | DAKO EnVision+ System, anti‐mouse | Novusbio |
| Smoothelin, R4A | Smooth |
Mouse IgG1 |
Not specified | Cytoskeleton (intermediate filament) | Tris‐EDTA (pH 9.2) | No | 1:300 | DAKO EnVision+ System, anti‐mouse | Thermo Fisher Scientific |
| Tropomyosin, F‐6 | Tropo | Mouse IgG2b | Not specified | Cytoskeleton (actin binding) | Tris‐EDTA (pH 9.2) | No | 1:8000 |
|
Santa Cruz Biotechnology |
| Myosin light chain kinase/Telokin A01697‐2 | Telokin | Rabbit IgG | Not specified | Cytoskeleton (myosin binding) | No retrieval | No | 1:1000 | DAKO EnVision+ System, anti‐rabbit | Bosterbio |
| Paxillin, 5H11 | Pax |
Mouse IgG1 |
Protein G |
Cytoplasm > cytoskeleton (focal adhesion protein) |
Tris‐EDTA (pH 9.2) | No | 1:500 | DAKO EnVision+ System, anti‐mouse | Invitrogen |
| Vinculin, hVIN‐1 | Vinc |
Mouse IgG1 |
Unpurified | Cytoskeleton (actin binding) | Citrate (pH 6) | No | 1:200 000 | DAKO EnVision+ System, anti‐mouse | Novusbio |
| Collagen‐I, MBS502155 | Col‐I | Rabbit IgG | Affinity chromatography | ECM | Citrate (pH 6), Tris‐EDTA (pH 9.2), and pepsin | No | No specific signal | DAKO EnVision+ System, anti‐rabbit | MyBioScource |
| Collagen‐I, C7510‐17K | Col‐I | Goat IgG | Affinity chromatography | ECM | Citrate (pH 6), Tris‐EDTA (pH 9.2), and pepsin | No | No specific signal | Donkey anti‐goat IgG‐HRP, AB97120 (Abcam) | USBIO |
| Procollagen‐I, PC8‐7 | Procol‐I | Mouse IgG1 | Not specified | Secreted | Citrate (pH 6) and Tris/EDTA (pH 9.2) | No | No specific signal | DAKO EnVision+ System, anti‐mouse | Abnova |
| Procollagen‐I, M58 | Procol‐I | Rat IgG1 | Affinity chromatography | Secreted | Citrate (pH 6) and Tris‐EDTA (pH 9.2) | No | No specific signal | Goat anti‐rat IgG‐Biotine, BA‐9401 (Vector) | Chemicon International |
| Prolyl 4‐hydroxylase β, 3‐2B12 | P4HB | Mouse IgG1 | Affinity chromatography | Endoplasmic reticulum lumen, cell membrane | Citrate (pH 6) | No | 1:4000 | DAKO EnVision+ System, anti‐mouse | Acris Antibodies |
| Osteopontin, AF1433 | OPN | Polyclonal goat IgG | Antigen affinity purified | ECM | Tris‐EDTA (pH 9.2) | No | 1:100 | Donkey anti‐goat IgG‐HRP, AB97120 (Abcam) | R&D Systems |
| Fibronectin, FBN11 | FBN | Mouse IgG1 | Protein G | ECM | Tris‐EDTA (pH 9.2) | No | 1:900 | DAKO EnVision+ System, anti‐mouse | Thermofisher |
| Laminin, ab11575 | Lam | Polyclonal rabbit IgG | Immunogen affinity purified | ECM | Tris‐EDTA (pH 9.2) | No | 1:200 | DAKO EnVision+ System, anti‐rabbit | Abcam |
| Cellular retinoid‐binding protein‐I, B8 | CRBP‐I | Mouse IgG1 | Not specified | Cytoplasm | Citrate (pH 6) and Tris‐EDTA (pH 9.2) | No | No signal | DAKO EnVision+ System, anti‐mouse | Santa Cruz Biotechnology |
| Platelet‐derived growth factor receptor α, ab61219 | PDGF‐α | Rabbit IgG | Immunogen affinity purified | Cell membrane | Citrate (pH 6) and Tris‐EDTA (pH 9.2) | No | A specific signal (nuclear) | DAKO EnVision+ System, anti‐rabbit | Abcam |
| Phosphorylated nuclear factor‐κB p105, 178F3 | NF‐κB | Rabbit IgG | Not specified | Nucleus | Citrate (pH 6), Tris‐EDTA (pH 9.2), and pepsin | No | No specific signal | DAKO EnVision+ System, anti‐rabbit | Cell Signaling Technology |
| Phosphorylated nuclear factor‐κB p65, MCFA30 | NFκB | Mouse IgG1 | Antigen affinity purified | Nucleus | Citrate (pH 6), Tris‐EDTA (pH 9.2), and pepsin | No | No specific signal | DAKO EnVision+ System, anti‐mouse | eBioscience |
| Interleukin 6, NYRhIL6 | IL‐6 | Mouse IgG2a | Not specified | Secreted |
Citrate (pH 6) |
Yes | 1:300 | DAKO EnVision+ System, anti‐mouse | Santa Cruz Biotechnology |
| Monocyte chemoattractant protein‐I, 23002 | MCP‐I | Mouse IgG2b | Protein A or G | Secreted | Citrate (pH 6), Tris‐EDTA (pH 9.2), and pepsin | No | No signal | DAKO EnVision+ System, anti‐mouse | R&D Systems |
| Interleukin 8, bs0708R | IL‐8 | Rabbit IgG | Protein A | Secreted | Tris‐EDTA (pH 9.2) | No | 1:600 | DAKO EnVision+ System, anti‐rabbit | Bioss |
| CD31, JC70A | CD31 | Mouse IgG1 | Tissue culture supernatant | Cell membrane | Tris‐EDTA (pH 9.2) | No | 1:1000 | MACH‐2 Biocare Medical, anti‐mouse | Dako |
| CD45, HI30 | CD45 | Mouse IgG1 | Affinity chromatography | Cell membrane | Tris‐EDTA (pH 9.2) | No | 1:2000 | MACH‐2 Biocare Medical, anti‐mouse | Biolegend |
| CD34, M1636 | CD34 | Mouse IgG1 | Affinity chromatography | Cell membrane |
Citrate (pH 6) |
Yes | 1:1 000 000 | MACH‐2 Biocare Medical, anti‐mouse | Sanquin |
| CD4, H‐370 | CD4 | Rabbit IgG | Not specified | Cell membrane | Tris‐EDTA (pH 9.2) | No | 1:800 | MACH‐2 Biocare Medical, anti‐rabbit | Santa Cruz Biotechnology |
| CD8, C8/144B | CD8 | Mouse IgG1 | Not specified | Cell membrane | Tris‐EDTA (pH 9.2) | No | 1:200 | MACH‐2 Biocare Medical, anti‐mouse | Dako |
|
Rabbit IgG Ref x0903 Lot 20011861 |
Dako | ||||||||
|
Rabbit IgG Ref I5006 Lot SLBK1653V |
Sigma‐Aldrich | ||||||||
|
Mouse IgG Ref x0931 Lot 20023783 |
Dako | ||||||||
|
Rabbit serum Ref x0902 |
Dako | ||||||||
|
Mouse serum Ref x0910 |
Dako |
CD indicates cluster of differentiation; and ECM, extracellular membrane.