Empty vector and rLOX-PP expressing PC3 and DU145 cells were cultured, extracted into non-denaturing RIPA cell lysis buffer and incubated with agarose bead-immobilized Non-immune IgG or anti-c-Myc tag IgG overnight at 4°C to pull down rLOX-PP. Non-immune IgG (#31903, Thermo Scientific, Waltham, MA) was coupled to agarose using AminoLink Immobilization Plus Trial kit (#20394, Thermo Scientific, Waltham, MA) according to the manufacturer’s instructions. For pull-down assays, beads were washed, and then eluted according to kit instructions (ProFound c-Myc Tag IP/Co-IP Kit; Thermo Scientific, Waltham, MA). Eluted samples were analyzed by Western blotting on denaturing SDS PAGE probed with an anti-ATM, anti-ATR, Rad50, anti-NBS1 and anti-MRE11 antibodies (Cell Signaling, Danvers, MA) and with anti-rLOX-PP antibody (61 (link)). Aliquots (5%) of initial extracts taken before the immunoprecipitation were analyzed by Western blot on the same gels.
Aminolink immobilization plus trial kit
Manufactured by Thermo Fisher Scientific
The AminoLink Immobilization Plus Trial kit is a lab equipment product from Thermo Fisher Scientific. It is designed for the covalent immobilization of proteins and other biomolecules. The kit includes all the necessary reagents and components to perform the immobilization process.
Automatically generated - may contain errors
Lab products found in correlation
Anti atr, by Cell Signaling Technology (2 mentions)
Rad50, by Cell Signaling Technology (2 mentions)
Anti nbs1, by Cell Signaling Technology (2 mentions)
Anti mre11, by Cell Signaling Technology (2 mentions)
Profound c myc tag ip co ip kit, by Thermo Fisher Scientific (2 mentions)
Non immune igg, by Thermo Fisher Scientific (2 mentions)
2 protocols using aminolink immobilization plus trial kit
1
Identification of rLOX-PP Interactors
2
Identification of rLOX-PP Interactors
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