Cells either were harvested in phosphate-buffered saline and were then undergoing sonification-assisted cell lysis in RIPA (
radioimmunoprecipitation assay) buffer in order to gain protein lysates or were further processed for RNA isolation.
Treated cells were lysed using
radioimmunoprecipitation assay buffer (Sigma-Aldrich, USA) containing 1% protease inhibitor cocktail and 2% phenylmethanesulfonyl fluoride.
Intestinal or brain homogenates were lysed in ice-cold
radioimmunoprecipitation assay lysis buffer with 0.02 mM phenylmethanesulfonyl fluoride for 30 min and then ultrasonicated for 60-s intervals in an ice bath.
Proteins from insoluble pellets were then extracted using buffers with increasing solubilization strength: HST buffer (HS buffer containing 1% Triton X-100), RIPA (
radioimmunoprecipitation assay) buffer, and 2% sodium dodecyl sulfate (SDS).
Radioimmunoprecipitation assay for glutamic acid decarboxylase antibodies evaluated clinically with sera from patients with insulin-dependent diabetes mellitus.
Serologic tests for HTLV-I, including enzyme immuno-assay, Western blot, and
radioimmunoprecipitation assay, also detect HTLV-II through cross-reactivity.
For the in vivo study, tissue fragments were lysed in
radioimmunoprecipitation assay buffer supplemented with a cocktail of protease inhibitors.
The LV tissue was lysed in
radioimmunoprecipitation assay (RIPA) lysis buffer, and the total protein was extracted and detected with a BCA Protein Assay Kit (Thermo Fisher Scientific, MA, USA).
For Western blotting, the cells were collected and lysed in
radioimmunoprecipitation assay (RIPA) buffer with protease inhibitors on ice.