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Affinity Measurement Services for Cell Lysates

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Creative Biolabs has strong expertise and extensive experience in affinity and kinetics measurement with cell lysate. We can satisfy any specific demand with our label-free and high efficiency affinity Surface Plasmon Resonance (SPR) and Bio-Layer Interferometry (BLI) technologies.

Various methods have been developed to analyze the affinity of interested protein with other proteins and components in crude cell lysate. One classic method is affinity purification, which needs the purified protein of interest to carry out the followed binding analysis step. However, Creative Biolabs can provide label-free protein affinity detection in crude cell lysate based on SPR technique with anitbody arrays, without the need of highly-purified protein samples. (Figure 1, left paenel).

Figure 1. SPR sensorgram of a lysate prepared with E. coli which expresses 6xHis-tagged 14-3-3 protein, the sample was injected over a sensorchip with immobilized Ni2+ while the chip was regenerated with 0.3 M imidazol (left); Overview of reduction of non-specific adsorption from cell lysate with peptide SAMs (right). (Mol. Biotechnol., 2006) Figure 1. SPR sensorgram of a lysate prepared with E. coli which expresses 6xHis-tagged 14-3-3 protein, the sample was injected over a sensorchip with immobilized Ni2+ while the chip was regenerated with 0.3 M imidazol (left); Overview of reduction of non-specific adsorption from cell lysate with peptide SAMs (right). (Mol. Biotechnol., 2006)

There are two problems in affinify measurement with cell lysate: 1). Bulk of effort, which can be overcomed by adjusting the reflectivity of running buffer with that of the lysate; 2). Non-specific adsorption, which can be reduced by the hydrophilicity and low electrostatic charge of the surface chemistries (Figure 1, right panel). The optimized system enables high-throughput analysis of diverse proteins with minimal sample volume (200 μL). The antibody array on the gold chip was placed into an SPR instrument (Figure 2).

Figure 2. The surface chemistries of the fabricated antibody arrays. (Anal. Chem., 2005)Figure 2. The surface chemistries of the fabricated antibody arrays. (Anal. Chem., 2005)

This system is applicable for, but not limited to:


Reference:

A. Aube et al. (2013). Non-specific Adsorption of Crude Cell Lysate on Surface Plasmon Resonance Sensors. Langmuir, 29 (32): 10141-1048.
J. Borch and P. Reopstorff (2006). Combinations of SPR and MS for Characterization of Native and Recombinant Proteins in Cell Lysates. Mol. Biotechnol., 33: 179-190. 
M. Kyo et al. (2005). Label-Free Detection of Proteins in Crude Cell Lysate with Antibody Arrays by a Surface Plasmon Resonance Imaging Technique. Anal. Chem., 77 (22): 7115-7121.




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