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Flow cytometric detection method of coelomocyte phagocytic activity of Apostichopus japonicus

A technology of flow cytometry and detection method, which is applied in the field of flow cytometry, can solve the problems of application limitation, not being widely used, poor repeatability, etc., and achieve the effect of simple and fast operation, good repeatability and high accuracy

Inactive Publication Date: 2011-12-14
SHANDONG UNIV AT WEIHAI
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

Among them, the microscope inspection method is suitable for the morphological observation of cells and the qualitative study of the phagocytosis function of single cells through the observation and analysis of microscopes. However, due to the shortcomings of this technique such as strong subjectivity and poor repeatability, its application is limited to a certain extent.
Flow cytometry (Flow-cytometry, FCM) can use different fluorescent markers to study the phagocytosis of cells. It has the characteristics of high accuracy, good repeatability, and easy and fast operation. conventional means, but it has not been widely used in echinoderms

Method used

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  • Flow cytometric detection method of coelomocyte phagocytic activity of Apostichopus japonicus
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  • Flow cytometric detection method of coelomocyte phagocytic activity of Apostichopus japonicus

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Embodiment Construction

[0020] The present invention will be described in further detail below in conjunction with specific examples, but the present invention is not limited to the following examples.

[0021] Prepare a Tris-HCl buffer solution with a pH of 7.6 and a concentration of 0.068mol / L, then add 0.02mol / L of EGTA, 0.34mol / L of NaCl, and 0.018mol / L of KCl, and finally filter it with a 0.22μm filter. anticoagulant.

[0022] Prepare a Tris-HCl buffer solution with a pH of 7.6 and a concentration of 0.001mol / L, then add 0.8mol / L sucrose, 0.34mol / L NaCl, and 0.001mol / L EGTA, and finally filter it with a 0.22μm filter. Sucrose buffer.

[0023] Prepare a Tris-HCl buffer solution with a concentration of 0.01 mol / L at pH 7.6, then add 0.01 mol / L EGTA and 0.34 mol / L NaCl, and finally filter with a 0.22 μm filter to obtain an isotonic buffer solution.

[0024] Extract 2.5ml of body cavity fluid of normal and diseased sea cucumbers with the same volume as the anticoagulant with a sterile syringe, put...

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Abstract

The invention discloses a flow type cell detection method for phagocytic activity of coelomic cells of a stichopus japonicus, belonging to the technical field of halobios. The method comprises the following steps of: firstly separating and preparing phagocytic cells in coelomic fluid of the stichopus japonicus; labeling heat-inactivated yeast cells by using fluorescein isothiocyanate (FIFC); co-incubating the labeled yeast cells and the phagocytic cells of the stichopus japonicus; and finally measuring the phagocytic activity by using flow cytometry. The method has high accuracy and good repeatability and is simple and convenient and fast to operate, and a method basis is provided for researching the phagocytic function of the coelomic cells of the stichopus japonicus.

Description

technical field [0001] The invention relates to a flow cytometric detection method for the phagocytosis activity of japonicus coelomocytes, belonging to the technical field of marine organisms. Background technique [0002] Due to the increasing market demand for sea cucumbers year by year, the traditional fishing industry can no longer meet people's needs. The sea cucumber aquaculture industry has emerged as the times require. The rapid increase in aquaculture area and output has created considerable social and economic benefits for coastal areas. However, in recent years, diseases of sea cucumbers have been constantly occurring. Since the winter of 2003, large-scale epidemics of rotten skin and swollen mouths have occurred in the cultured sea cucumbers in Shandong and Liaoning coastal areas, causing a large number of deaths of more than 30%. Immunology research is a science that understands the characteristics of the body's immune response, thereby laying the foundation an...

Claims

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Application Information

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IPC IPC(8): G01N15/14
Inventor 刘洪展郑风荣屈佩孙修勤
Owner SHANDONG UNIV AT WEIHAI
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