Protein immunogold labeling method for cryoelectron microscope

Abstract

The invention provides a protein immunogold labeling method for a cryoelectron microscope. The protein immunogold labeling method comprises the following steps of (1) respectively pretreating a grid of the frozen electron microscope by utilizing a poly-L-lysine solution and PBS (Phosphate Buffer Solution); (2) putting the grid in a culture dish with the front surface facing upwards, dripping a sample onto the grid, standing, incubating, and eluting with PBS (Phosphate Buffer Solution); 3) adding a sealing agent into the culture dish, standing, incubating, and eluting with PBS; 4) adding a primary antibody, namely a specific antibody of the target protein, into the culture dish, standing, incubating, and eluting with PBS; 5) adding a secondary antibody, namely an immune gold antibody selected according to the species of the primary antibody, into the culture dish, standing, incubating, and eluting with PBS; and 6) taking out the grid, imaging by using a freezing electron microscope, andanalyzing image data to obtain a target protein structure. The method is advantaged in that an immunolabelled protein technology and a freezing electron microscope tomography technology are combined,so the target protein is observed in the freezing electron microscope imaging, and the structure is analyzed at the same time.

Description

technical field [0001] The invention relates to the technical field of immunolabelling, in particular to an immunogold-labeled protein method for cryo-electron microscopy. Background technique [0002] Immunolabeling protein technology is widely used in clinical and scientific research, such as the analysis of pathological sections at room temperature. This technology plays a role in labeling proteins by specifically binding protein antibodies to proteins after incubation, but conventional immunolabeling methods can only be used to distinguish and trace proteins, that is, to prove that the protein is contained in the sample and cannot be seen The specific structure of the protein. [0003] At present, the commonly used means of analyzing protein structure is cryo-electron microscopy tomography, which is a microscopic technique that uses transmission electron microscopy at low temperature to observe the structure of samples. Reconstruct the three-dimensional structure of th...

AI Technical Summary

Problems solved by technology

However, the disadvantage of cryo-electron microscopy is that it can only display the three-dimensional structure of the protein and cannot label the protein.

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