Anti rat hrp

The following primary antibodies were used for flow cytometry: anti-MICA (clone 159227; R&D Systems), anti-MICB (clone 236511, R&D Systems), anti-ULBP1 (clone 170818; R&D Systems), anti-ULBP2/5/6 (clone 165903; R&D Systems) and anti-ULBP3 (clone 166514, R&D Systems).
The following primary antibodies were used for immunofluorescence: anti-PDI (ab3672, Abcam), anti-FLAG tag (Clone L5, Biolegend) and anti-MICA (clone 159227, R&D Systems).
The following primary antibodies were used for western blotting: anti-MICA (Clone EPR6568, Abcam), anti-FLAG tag (Clone L5, Biolegend), anti-GAPDH (clone 6C5, Santa Cruz) and anti-vinculin (clone EPR8185, Abcam).
The following secondary antibodies were used: anti-mouse AlexaFluor 647, anti-mouse PE, anti-mouse biotin, anti-rabbit biotin, anti-rat biotin, anti-rabbit Cy3, anti-rat 488, streptavidin-AlexaFluor 647, streptavidin-horseradish peroxidase (HRP), anti-mouse-HRP, anti-rat-HRP and anti-rabbit-HRP, all purchased from Jackson Laboratories.

Frozen cell pellets were dissolved in Laemmlie-buffer (60 mM Tris-HCl, 10% glycerol, 2% SDS, 10% β-mercaptoethanol, 0.01% bromophenol blue; pH 6.8) (10 µl buffer per 100,000 cells) and subjected to sonication (3–5 s on ice with a UP50H sonicator equipped with an MS1 sonotrode) (Hielscher, Teltow, Germany). Samples were then heated to 89°C for 3 min, spun for 5 min at room temperature and the supernatants used for standard SDS-PAGE with 12% gels. Wet blotting was carried out in Mini Trans-Blot modules (Bio-Rad Laboratories) using nitrocellulose membranes and blotting buffer (20% v/v methanol, 25 mM Tris-HCl, 192 mM glycine, pH 8.6). The following antibodies were used for target detection: anti-ERK1/2 (Cell Signaling Technology, Frankfurt am Main, Germany; no. 9102), anti-ERK1/2 (Santa Cruz Biotechnology, Heidelberg, Germany; sc-94), anti-phospho-ERK1/2 (Cell Signaling Technology; no. 9101), anti-tubulin (Biozol, Eching, Germany; BZL03568). The secondary antibodies were from Jackson ImmunoResearch Laboratories (Newmarket, UK): anti-rat-HRP (112-036-062), anti-rabbit HRP (111-036-045). A freshly made solution of luminol (2.5 mM), p-coumaric acid (0.2 mM) and H₂O₂ (0.01%) in 100 mM Tris-HCl (pH 8.8) was used for chemiluminescent detection [29] (link).