Dawn mals detector

SEC-MALS analysis was performed using a DAWN MALS detector (Wyatt Technology) connected to an AKTA FPLC system (GE Healthsciences). CtsK, CtsK/12merNS2S6S (compound # 20) and CtsK/12merNS6S (compound # 24) complexes were prepared as described above and concentrated to ~2mg/ml, and 100 µl was resolved on Superdex 200 Increase SEC column using 20 mM NaOAc, 250 mM NaCl, pH 5.5. The MALS data were analyzed using ASTRA software (ver. 7.3.2.17).

SEC-MALS was carried out using a Superdex 200 gel filtration column on an AKTA fast liquid chromatography system (Cytiva Life Sciences) coupled with a DAWN MALS detector and an Optilab refractive index detector (Wyatt Technology, Santa Barbara, CA). Data for IC_1-260 was collected for protein samples at a concentration of 200 µM protein in a buffer composed of 50-mM sodium phosphate (pH 7.5), 50-mM sodium chloride, and 1-mM sodium azide. Data for IC_FL was collected for protein samples at a concentration of 30 µM in a buffer composed of 25-mM tris(hydroxymethyl)aminomethane hydrochloride (pH 7.4), 150-mM KCl, 5-mM β-mercaptoethanol, and 1-mM sodium azide. Molar mass and error analysis were determined using ASTRA v9, employing a Zimm light scattering model (Wyatt Technology).

SEC-MALS analysis was performed using a DAWN MALS detector (Wyatt Technology) connected to an AKTA FPLC system (GE Healthsciences). mTRAIL, mTRAIL/12mer complex and mTRAIL/18mer complex were prepared as describe above and concentrated to ~4 mg/ml, and 100 µl was resolved on Superdex 200 Increase SEC column using 25 mM HEPES, 150 mM NaCl, pH 7.1. The MALS data was analyzed using ASTRA software (ver. 7.3.2.17).

SEC-MALS was performed on an AKTApure FPLC (Cytiva) with a DAWN MALS detector (Wyatt). A total of 100 µg of ESAT-6 was buffer exchanged with a Zeba spin column immediately prior to each experiment and injected onto a Superdex 75 increase 10/300 column at a flow rate of 0.5 mL/min at 4 °C. The mobile phase was 10 mM Citrate, 300 mM NaCl, pH 4.5 or pH 7.5. Buffers were sterile filtered to 0.1 µm and degassed prior to use. Data analysis was performed using ASTRA 8.1.2.1 (Wyatt).

A 50 μL aliquot of 0.5 mg/mL S. enterica WbaP in SMALP was injected onto a WTC-030 fused silica column pre-equilibrated with HEPES buffered saline pH 7.5 (HBS). Eluate was flowed through a DAWN MALS detector (Wyatt) and an Optilab differential refractive index detector (Wyatt). Data were analyzed using ASTRA software (Wyatt), and the system was pre-calibrated with a BSA standard.