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| Comparison of Slide-preparing Methods of Chicken Bone Marrow Dendritic Cells by Indirect Immunofluorescence |
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| DOI：10.16768/j.issn.1004-874X.2020.10.016 |
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| Abstract: |
| 【Objective】In order to improve the laboratory techniques and methods of dendritic cells（DC）, two slide-preparing methods（cell climbing and cell dropping）of DC from chicken bone marrow by indirect immunofluorescence was compared and analyzed.【Method】Using chicken embryos as test material, the immature bone marrow cells were isolated and obtained from chicken in vitro, and then were activated by Newcastle disease virus（NDV） lastota strain. The expressions of endogenous protein ROBO2（Roundabout） and exogenous protein NDV were monitored. Chicken NDV antibody and mouse ROBO2 monoclonal antibody were hatched together and marked with the combination of FITC. Fluorescence imaging of dendritic cells was examined by laser scanning confocal microscopy（LSCM）, and the difference between cell climbing and cell dropping was analyzed variously.【Result】The cell dropping process was time- consuming and labor intensive, while the cell climbing process was characterized by easy operation and reduced time. NDV and ROBO2 of these two methods showed highly significant differences in single positive and negative and significant influorescence reactivity differences in double positive.【Conclusion】The immunoreactivity of the protein was better with the cell dropping, which was suitable for labeling target and subcellular localization. The morphological and protein characteristics were always
maintained by the cell climbing,which was beneficial to the fluorescence overlap of the two antibodies and to make the overall color rendering stronger. |
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