Method for separating babylonia areolata granular cells and clear cells by Percoll serving as medium

A technology of snail snails and granule cells, applied in cell dissociation methods, biochemical equipment and methods, animal cells, etc., can solve the problems of poor understanding of the immune function of shellfish blood cells, etc., to reduce the probability of pollution, good application Foreground, simple effect

Pending Publication Date: 2019-08-16
HENAN NORMAL UNIV
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Problems solved by technology

At present, it is generally believed that the two types of blood cells in shellfish have been accepted by most people: granule cells and clear cells. The applicant has initially clearly divided blood cells into granule cells and clear cells through the study of the blood cell morphology of Fangban Dongfeng snails. However, shellfish blood cells Immune function remains poorly understood

Method used

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  • Method for separating babylonia areolata granular cells and clear cells by Percoll serving as medium
  • Method for separating babylonia areolata granular cells and clear cells by Percoll serving as medium
  • Method for separating babylonia areolata granular cells and clear cells by Percoll serving as medium

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Embodiment

[0024] 1. Preparation of Density-Optimized Percoll Solution

[0025] The density of the Percoll stock solution is 1.13g / mL; use the anticoagulant ZA solution to prepare Percoll solutions with a volume fraction of 38% and a volume fraction of 35%.

[0026] 2. Blood cell concentration adjustment

[0027] Suspend the hemocytes of the snail and use the anticoagulant ZA solution to adjust the blood cell density of the snail to (2–5)×10 5 Cells / mL to obtain the blood cell suspension of the snails.

[0028] 3. Ratio of separating liquid and sample volume

[0029]Using a 15mL centrifuge tube, add 3mL of Percoll solution with a volume fraction of 38% and 2mL of blood cell suspension of P.

[0030] 4. Two-step centrifugation

[0031] Centrifuge at 1300g at 4°C for 45min. The solution is divided into three layers: the upper layer, the lower layer and the interface layer. Separately pack the upper layer solution, the lower layer solution, and the interface solution between the upper a...

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Abstract

The invention belongs to the technical field of babylonia areolata hemocyte separation and discloses a method for separating babylonia areolata granular cells and clear cells by Percoll serving as medium. The technical scheme is characterized in that the method includes steps: taking a 15mL centrifuge tube, adding 3mL Percoll solution in volume fraction of 38% and 2mL of babylonia areolata hemocyte suspension, and centrifuging for 45min under conditions of 1300g and 4 DEG C; after solution is layered into upper and lower layers and an interface layer between the upper and lower layers, takinganother new 15mL centrifuge tube, adding 3mL Percoll solution in volume fraction of 35% and 1mL of interface solution of the upper and lower layers, centrifuging for 45min under conditions of 1300g and 4 DEG C, and collecting upper-layer solution and lower-layer solution respectively to finally realize separation of babylonia areolata granular cells and clear cells. A separation process is simple,convenient, easy in operation, high in specificity and low in cost.

Description

technical field [0001] The invention belongs to the technical field of blood cell sorting of Dongfeng snails, and in particular relates to a method for separating granule cells and clear cells of Dongfeng snails by using Percoll as a medium. Background technique [0002] In vertebrates, innate and adaptive immune functions protect the host from infection. However, invertebrates lack an adaptive immune system, and innate immunity plays a crucial role in the response to infectious pathogens. The innate defense system of invertebrates, including gastropods, is divided into humoral and cellular immunity. The humoral immune response in insects is currently well understood. However, cellular immune responses have not been extensively explored. Cellular immune responses in invertebrates mediated by circulating blood cells play a key role in immobilizing or destroying invading pathogens. [0003] It is generally believed that shellfish do not have acquired immunity based on lymp...

Claims

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Application Information

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IPC IPC(8): C12N5/07
CPCC12N5/0601C12N2509/00
Inventor 狄桂兰李艳菲孔祥会李莉裴超赵贤亮张杰赵燕静
Owner HENAN NORMAL UNIV
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