echemi logo
Product
  • Product
  • Supplier
  • Inquiry
    Home > Research group of Professor Su bin of Zhejiang University: electrochemiluminescence microscopy for cell matrix adhesion and cell mass migration imaging of living cells

    Research group of Professor Su bin of Zhejiang University: electrochemiluminescence microscopy for cell matrix adhesion and cell mass migration imaging of living cells

    • Last Update: 2019-11-15
    • Source: Internet
    • Author: User
    Search more information of high quality chemicals, good prices and reliable suppliers, visit www.echemi.com
    Lead electrochemiluminescence is a kind of chemiluminescence triggered by electrochemical method, which has two obvious advantages: strong spatiotemporal controllability and low background signal The electrochemiluminescence microscope is obtained by using electrochemiluminescence as the light source of the microscope As a powerful tool to study micro and nano scale objects, electrochemiluminescence microscopy has realized imaging from micro and nano particles to single cells Recently, the research group of Professor Su bin, Department of chemistry, Zhejiang University has made new progress in this field (angel Chem Int ed., 2019, DOI: 10.1002/anie.201911190) Brief introduction of Professor Su Bin's research group Professor Su Bin's research group focuses on the research direction of electrochemiluminescence methods and technologies, micro nano scale molecular separation and analysis, and carries out the basic and Application Research on spatiotemporal resolution electrochemiluminescence, electrochemiluminescence immune / molecular diagnosis, micro interface biomimetic sensor analysis, etc Prof Su bin, Professor of the Department of chemistry, Zhejiang University, director of the Institute of analytical chemistry, Zhejiang University Mainly engaged in the basic and applied research of Interface Electrochemistry, photoelectrochemical measurement and imaging method / technology, micro and nano scale molecular separation and analysis, fingerprint identification and trace inspection and analysis So far, more than 130 papers have been published, including 17 If > 10 (including 7 J am Chem SOC., 6 Angel Chem Int ed., 2 ACS Nano), more than 50 if > 5, and 9 authorized Chinese invention patents More than 30 graduate students have been guided, including one who has won the academic newcomer award of doctoral degree from the Ministry of education, one who has won Zhu Kezhen scholarship from Zhejiang University, and four who have won the national scholarship Cutting edge research results: when the electrochemiluminescence microscope is used for cell matrix adhesion and cell collective migration imaging of living cells to culture adherent cells, cell matrix adhesion is formed between cells and the basement (Figure 1) Cell matrix adhesion is a complex transmembrane structure composed of many proteins It connects with cytoskeleton in the cell, recognizes specific adhesion sites outside the cell, and anchors the cell to extracellular matrix But the function of cell adhesion is not only to fix cells, but also to play an important role in cell migration, wound healing, cancer occurrence and metastasis At present, cell matrix adhesion imaging methods include scanning / transmission electron microscopy, reflection interference microscopy, fluorescence and surface plasmon resonance microscopy Although these methods have helped biologists to solve some problems, they also have limitations, such as the operation of electron microscopy needs to fix cells, fluorescence imaging depends on markers and so on In this research group, a single cell electrochemiluminescence imaging system was constructed to realize the cell matrix adhesion imaging without markers Fig 1 The principle of microimaging of electrochemiluminescence cell adhesion (source: angel Chem Int ed., 2019, DOI: 10.1002/anie.201911190) when imaging ECL cells, cells need to be pre incubated on ITO electrode modified by SNM The electrode can enhance the electrochemiluminescence of Ru (bpy) 32 + / CO reagent by two orders of magnitude In the experiment, 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES) was selected as a co reagent, which is also a biological buffer, and can effectively control the pH of the solution in the open cell culture environment Ru (bpy) 32 + and HEPES were added to PBS solution, and a suitable potential was applied to the electrode The reaction as shown in formula 1-6 occurred on the electrode surface without cells, and photons were released The light-emitting sites were bright areas in the image, in which B (buffer) and B · respectively represented HEPES and its free radical intermediates, and P was the product of B · homogeneous oxidation reaction When the cells adhere to the surface of SNM, they inhibit the diffusion of Ru (bpy) 32 + and HEPES to the surface of ITO electrode The light emission is weak and appears as a gray area That is to say, the reverse electrochemiluminescence image of cell adhesion is obtained Fig 2 shows the transmission light field of PC12 cells and the corresponding ECL image In the two imaging modes, the cell contour basically corresponds, but the difference is also obvious In the ECL image, the blurry cell structure (shown by the white arrow) in the open field mode can be clearly observed, such as cell protrusion (Fig 2D), pseudopodia (Fig 2e), curling (Fig 2F right side) and some short and orderly cell structures similar to the adhesion spot (Fig 2F left side) In addition, not all areas in the cell ECL image are dark (as shown by the white circle), and the gray value is close to the bright background value of the cell-free area This is because the bottom of the cell will not form adhesive structure, and the luminescence of the non adhesive area will not be inhibited Fig 2 Open field (A-C) and corresponding ECL image (d-f) of PC12 living cells (source: angel Chem Int ed., 2019, DOI: 10.1002/anie.201911190) pancreatin can digest cell adhesion and separate cells from the basement The dynamic changes of adhesion were studied by ECL imaging using trypsin digestion as a model Figure 3 compares the open field and ECL images of a single PC12 cell After adding trypsin for three minutes, it was observed that the upper part of cells retracted and became round in the open field (Fig 3b), the loose adhesive structure in the upper part of cells disappeared in the ECL image, and the compact structure in the lower part basically remained unchanged (Fig 3e) After six minutes, the upper part of cells in the open field continued to become round (Fig 3C), and ECL imaging showed that the adhesion area gradually decreased (Fig 3f) Cells are in a uniform trypsin solution, and are not interfered by other external forces Therefore, the results obtained under this condition can be used to characterize the adhesion strength of cells The adhesive strength of the upper part of the cell is low and easy to be digested, while that of the lower part is high and difficult to be digested In order to study the relationship between cell digestion rate and adhesion morphology, the author defined the digestion rate as the change of adhesion area of single cell in unit time For cells with the same adhesive area, the higher the digestion rate, the lower the adhesive strength Figure 3g-i shows the relationship between PC12 cell digestion rate and cell adhesion area, perimeter and ferret's diameter, respectively The results showed that there was a positive correlation between the digestion rate and the three parameters, and the Pearson correlation coefficient (PCC) was 0.949, 0.798 and 0.782, respectively, that is, the highest correlation between digestion rate and adhesion area The larger the adhesive area is, the larger the contact area between the adhesive protein and trypsin is, the larger the reaction cross section is, and the higher the reaction rate is In addition, the average digestion rate of Huh-7 cells was lower than that of PC12 cells, and the gray value of ECL image of Huh-7 cells was lower than that of PC12 cells The lower gray level of ECL image may be attributed to the higher adhesive strength Therefore, in the absence of trypsin, the adhesive strength can be roughly determined by the ECL gray scale Figure 3 Open field (A-C) and ECL image (d-f) of single PC12 cell digested by trypsin: (a, d) 0 min, (B, e) 3 min, (C, f) 6 min (G-I) the statistical relationship between single cell trypsin digestion speed and cell adhesion area (g), perimeter (H) and ferret's diameter (I) (source: angel Chem Int ed., 2019, DOI: 10.1002/anie.201911190) cell adhesion plays a key role in cell collective migration People have a clear understanding of the adhesion changes of single cells in the process of migration However, in the process of collective cell migration, how cell groups perceive the direction of migration, and what role "leader" cells and "follower" cells play, have not been determined The following authors used ECL imaging for the analysis of cell mass migration and adhesion Compared with the cell open field and ECL images at different times in the scratch experiment, the cells in the open field images taken at 0 h and 2 h after cell migration were all disordered (FIG 4A and b), while in the ECL image, the cell adhesion showed a significant migration tendency towards the scratch direction (Fig 4D) In order to describe the directional migration of cells, the author analyzed the vector field distribution of the bright field and ECL image, as shown by the red lines in Fig 4C and D and the black arrows in E and F In Figure 4G and h, the vector field distribution is displayed in the form of color chart, and the difference of vector distribution at two times can be observed clearly: the distribution boundary of vector field in G chart is clear, parallel to the scratch boundary, and the vector field at the same position in H chart is more perpendicular to the scratch direction This result can not be obtained by the analysis of the bright field image Similarly, the author analyzed the vector field of ECL image obtained by cell migration for 4-12h, and the statistical results are shown in Figure 5 Figure 4 Open field (a, b) and ECL images (C, d) of monolayer PC12 cells in cell scratch experiment The red lines in figures C and d represent the local cell orientation, which are shown in the form of directional arrows and color pictures in figures e, F, G and h, respectively The first image (a, C, e, g) was collected immediately after scratch, and the second image (B, D, F, H) was collected 2 hours after cell migration (source: angelw Chem Int ed., 2019, DOI: 10.1002/anie.201911190) the vector defined in the range of ± 45 ° is "directional vector", which represents the migration tendency of cells to scratches As shown in Fig 5a, when migration starts, the proportion of directivity vector is higher near the scratch and a platform appears The farther away from the scratch, the lower the proportion of directivity vector With the extension of migration time, the scope of the platform becomes larger and larger Defines the range where the directivity vector scale is greater than 80% as "directivity distance" As shown in Fig 5b, the distance increases with time and is far greater than the distance of cell migration For example, when migrating for 12 hours, the directional distance is about 350 m, while the migration distance is only 100 m This shows that the migration direction is determined not only by the cells migrating at the scratch edge, but also by the cells far away from the scratch This result emphasizes the role of "follower" cells in migration, which are not passive followers Figure 5 (a) directivity vector proportion varies with distance from scratch; (b) directivity distance (black) and migration distance (obtained from scratch experiment, blue) varies with time (source: angelw Chem Int ed., 2019, Doi: 10.1002/anie.201911190) in this work, a cell matrix adhesion free electrochemiluminescence imaging system was constructed The dynamic changes of subcellular level cell adhesion and the relationship between single cell level adhesion strength and adhesion morphology were studied, and the directional migration cells in collective migration were imaged and analyzed The research results were recently published in Angewandte Chemie International Edition (DOI: 10.1002 / anie 201911190) The corresponding author of the paper is Professor Su bin, Department of chemistry, Zhejiang University The first author is Professor Su Bin's doctoral student Ding Hao in the research group, and the co first author is Dr Guo Weiliang, postdoctoral student in Zhejiang University This work has been supported by NSFC and Natural Science Foundation of Zhejiang Province Thank you! Nowadays, people and scientific research have been paid more and more attention in the economic life China has ushered in the "node of science and technology explosion" Behind the progress of science and technology is the work of countless scientists In the field of chemistry, in the context of the pursuit of innovation driven, international cooperation has been strengthened, the influence of Returned Scholars in the field of R & D has become increasingly prominent, and many excellent research groups have emerged in China Therefore, CBG information adopts 1 + X reporting mechanism, CBG information website, chembeangoapp, chembeango official microblog, CBG information micro
    This article is an English version of an article which is originally in the Chinese language on echemi.com and is provided for information purposes only. This website makes no representation or warranty of any kind, either expressed or implied, as to the accuracy, completeness ownership or reliability of the article or any translations thereof. If you have any concerns or complaints relating to the article, please send an email, providing a detailed description of the concern or complaint, to service@echemi.com. A staff member will contact you within 5 working days. Once verified, infringing content will be removed immediately.

    Contact Us

    The source of this page with content of products and services is from Internet, which doesn't represent ECHEMI's opinion. If you have any queries, please write to service@echemi.com. It will be replied within 5 days.

    Moreover, if you find any instances of plagiarism from the page, please send email to service@echemi.com with relevant evidence.