蛋白组学 - 生物分子相互作用工作站Fortebio

来自 : biocore.sjtu.edu.cn/visitor/vi 发布时间：2021-03-25

仪器介绍(Apparatus Introduction)

主要性能指标
检测技术：基于光纤传感器的生物膜层光学干涉技术（BLI）
样品体积：终稀释液180-220μl，非破坏性检测
样品平板轨道运动：静止，或100-1500rpm
基线噪音：≤3 pm (RMS)
分析温度范围：室温+ 4°C至47℃，增量单位1℃
自动处理样品量：8道光纤生物传感器，一次同时做8个检测，最多自动完成96个检测
分析时间：8个样品分析时间 2min,96个样品分析时间 32min
定量范围：200rpm：0. 5-2000μg/mL 1000rpm：0. 05-100μg/mL
仪器主要用途
可分析样品种类：低分子量分析物，多肽，蛋白质；适合液体环境：含血清培养基，含
DMSO缓冲液，胞质悬液，未处理的细胞培养上清，粗制的细胞裂解液
动力学和亲和力分析(kobs, ka, kd, KD)
蛋白质定量
注意事项
1. 用SA/SSA传感器，蛋白首先要生物素化，该反应室温2小时
2. 蛋白浓度控制在1 mg/ml左右，一个传感器固化蛋白量约为10-50 μg
3. 做好对照实验
4. 生物素化会封闭Lys，影响Lys参与的结合
5. 最终计算Kd值需要浓度信息

Specification

Detection Technology: Bio-Layer Interferometry (BLI)
Sample Volume: 180–220 μL/well Nondestructive testing
Orbital Flow Capacity: Static or 100–1500 rpm
Baseline Noise: ≤3 pm (RMS)
Analysis Temperature Range: (Ambient + 4°C) – 40°C, 1°C increments
Throughput: Up to 8 assays in parallel; up to 96 assays per 96-well plate
Analysis Time：hIgG quantitation in 2 minutes for 8 samples, ≤32 minutes for 96
samples
Quantitation Range for hIgG：0.5 μg/mL to 2000 μg/mL at 200 rpm, 0.05 μg/mL
to 100 μg/mL at 1000 rpm
Application
Sample Types: Proteins, antibodies, peptides, media containing serum, buffers
containing DMSO, periplasmic fractions, untreated cell culture supernatants, and
crude cell lysates
Kinetic and affinity analysis(kobs, ka, kd, KD)
Protein quantification

Attention:
1. Protein must be biotinylated for SA/SSA biosensor. The reaction process for 2 h at ambient.
2. The concentration of protein is 1 mg/ml, and 10-50 μg protein would be needed for one biosensor.
3. Control experiment is necessary.
4. Lys would be sealed by the biotinylation, and influenced the result of the experiment.
5. The calculation of Kd need the information of the concentration.
文章 文献：
1. L. J. Wang, H.Z. Tang, H. Yu, Y. X. Yao, P. Xu, An unusual repressor controls the expression of a crucial nicotine-degrading gene cluster in Pseudomonas putida S16, Mol. Microbio. 2014, 91, 1252-1269.
2. H. Guo, Q. Zhu, X. Yu, S. B. Merugu, H. B. Mangukiya, N. Smith, Z. Li, B. Zhang, H. Negi, R. Rong, K. Cheng, Z. Wu, D. Li, Tumor-secreted anterior gradient-2 binds to VEGF and FGF2 and enhances their activities by promoting their homodimerization, Oncogene, 2017, 1-12.