Protean i12 isoelectric focusing system

The tissue samples were homogenized by using MediMachine and total protein isolation from the homogenized samples were isolated by lysing cells in lysis buffer (7 mol urea, 2 mol thiourea, 4% 3-((3-cholamidopropyl)dimethylammonio)-1-propanesulfonate solvent, 30 mM Tris–HCl, pH 8.5) and subsequent sonication (65% power, 20 seconds × 3 times). The concentrations of the protein samples were quantified by using Bradford assay. Two hundred fifty micrograms of total protein was used for 2-dimensional analysis. Isoelectric point separation was performed on 3 to 10 pH strips by using Protean i12 IsoElectric Focusing system (Bio-Rad) and subsequent gel electrophoresis was performed by using 12% Sodyum Dodesil Sülfat Gel. PDQuest Advanced 2-Dimensinal Analysis Software (BIO-RAD) was used for identification of differentially expressed proteins (Figs. 4 and 5).

Native Equ c 1 purified from the Breed/Gender Mix horse hair extract was prepared (2‐D Clean‐Up Kit, GE Healthcare) for two‐dimensional (2D)‐gel electrophoresis that was performed with immobilized pH gradient strips using the PROTEAN i12 isoelectric focusing system (Bio‐Rad Laboratories) as previously described.
¹⁷ (link) Gels were stained with SYPRO Ruby protein gel stain (Invitrogen, Thermo Fisher Scientific) according to the manufacturer's instructions. Images were recorded using the Amersham typhoon gel and blot imaging system (Amersham Typhoon 5, Cytiva).