Experimental Isoelectric Point (pI)

General Biophysical Characteristics

Isoelectric Point (pI)

unitless

UV Detection

None

None

None

None

280 nm

None

The pI was determined using the iCE3 system (Protein Simple) equipped with an UV detector at 280 nm and an Alcoot 720N autosampler. The separation was conducted using a 50 mm long, 100 µm (I.D.) fluorocarbon coated capillary (Protein Simple). The catholyte consisted of 0.1 M NaOH in 0.1% methyl cellulose and the anolyte of 0.08 M phosphoric acid in 0.1% methyl cellulose. According to the manufacturer´s protocol, protein samples were mixed with 1% methyl cellulose (0.4% final concentration), 5 µL of pharmalytes (GE Healthcare; final concentration 4%), 0.5 µl of low pI marker (pH 4.22) and 0.5 µL high pI marker (pH 9.22; final concentration: 1%) and sterile water until a final volume of 120 µL and final protein concentration of 0.2-0.5 mg/mL was reached. Samples were focused for 1 min at 1500 V and mobilized for 10 min at 3000 V with detection at 280 nm. Data processing was performed using Chrom Perfect (version 6.0.4).