Method for detecting PBMC activity under in-vitro 3D culture condition by using resazurin reagent

A culture condition, resazurin technology, applied in biochemical equipment and methods, measuring devices, and microbial determination/inspection, etc., can solve problems such as limited cell yield, and achieve the effects of convenient operation and high detection sensitivity

Pending Publication Date: 2020-06-19
郑州鲲鹏健康科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, adherent cells have the property of contact inhibition. Once the cells form a monolayer, the growth is inhibited and the cell yield is limited.

Method used

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  • Method for detecting PBMC activity under in-vitro 3D culture condition by using resazurin reagent
  • Method for detecting PBMC activity under in-vitro 3D culture condition by using resazurin reagent
  • Method for detecting PBMC activity under in-vitro 3D culture condition by using resazurin reagent

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Experimental program
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Effect test

Embodiment 1

[0025] A method for detecting PBMC activity under in vitro 3D culture conditions using a resazurin reagent, the specific steps are as follows:

[0026] Step 1, preparation of resazurin mother liquor: take 1 g of resazurin sodium and add 200 mL of three-distilled water to prepare a solution with a final concentration of 0.5%, and use a disposable filter with a pore size of 0.22 μm to sterilize the resazurin solution. Then put the sterilized resazurin solution into a 4°C refrigerator and store it for later use;

[0027] Step 2, preparation of resazurin detection reagent: according to the needs of the experiment, take an appropriate amount of resazurin mother solution, add a certain amount of three-distilled water, and prepare a resazurin solution with a final concentration of 0.15%, which is the resazurin detection Reagent;

[0028] Step 3: Determination of PBMC activity under in vitro 3D culture conditions: according to the ratio of the volume of the 3D agarose hydrogel medium...

Embodiment 2

[0031] A method for detecting PBMC activity under in vitro 3D culture conditions using a resazurin reagent, the specific steps are as follows:

[0032] Step 1, preparation of resazurin mother liquor: take 1 g of resazurin sodium and add 200 mL of three-distilled water to prepare a solution with a final concentration of 0.5%, and use a disposable filter with a pore size of 0.22 μm to sterilize the resazurin solution. Then put the sterilized resazurin solution into a 4°C refrigerator and store it for later use;

[0033] Step 2, preparation of resazurin detection reagent: according to the needs of the experiment, take an appropriate amount of resazurin mother solution, add a certain amount of three-distilled water, and prepare a resazurin solution with a final concentration of 0.17%, which is the resazurin detection Reagent;

[0034]Step 3: Determination of PBMC activity under in vitro 3D culture conditions: according to the ratio of the volume of the 3D methylcellulose hydrogel...

Embodiment 3

[0037] A method for detecting PBMC activity under in vitro 3D culture conditions using a resazurin reagent, the specific steps are as follows:

[0038] Step 1, preparation of resazurin mother liquor: take 1 g of resazurin sodium and add 200 mL of three-distilled water to prepare a solution with a final concentration of 0.5%, and use a disposable filter with a pore size of 0.22 μm to sterilize the resazurin solution. Then put the sterilized resazurin solution into a 4°C refrigerator and store it for later use;

[0039] Step 2, preparation of resazurin detection reagent: according to the needs of the experiment, take an appropriate amount of resazurin mother solution, add a certain amount of three-distilled water, and prepare a resazurin solution with a final concentration of 0.2%, which is the resazurin detection Reagent;

[0040] Step 3: Determination of PBMC activity under in vitro 3D culture conditions: according to the ratio of the volume of the 3D collagen hydrogel medium...

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Abstract

The invention discloses a method for detecting PBMC activity under an in-vitro 3D culture condition by using a resazurin reagent. The method comprises the following specific steps: step 1, preparing resazurin mother liquor; step 2, preparing a resazurin detection reagent; and step 3, determining the activity of PBMC under an in-vitro 3D culture condition. The standard resazurin solution which canbe produced on a large scale and is used for detecting the activity of human peripheral blood mononuclear cells after being cultured in an in-vitro 3D culture system in vitro is established; the reagent is high in cell activity detection sensitivity, convenient to operate, free of adverse effects on cells and free of pollution to the environment, and it is proved through actual tests that the reagent is suitable for detecting the activity of PBMC in a 3D culture system.

Description

technical field [0001] The invention relates to the technical field of in vitro diagnosis, in particular to a method for detecting PBMC activity under 3D culture conditions in vitro using a resazurin reagent. Background technique [0002] In vitro cell culture refers to the process of culturing organs, tissues or cells taken out of a living body in vitro by simulating the physiological conditions in the body, and allowing them to continue to survive, grow and reproduce. In vitro cell culture has become one of the basic techniques of modern biological research and is widely used in various aspects of modern biomedicine and biological science research. [0003] Currently, studies at the cell level in vitro are largely done under two-dimensional culture conditions. When cells grow under two-dimensional conditions, due to the inability to reproduce complex cell signals between cells and their substrates, cells gradually lose their original traits in vivo, and their morphology, ...

Claims

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Application Information

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IPC IPC(8): G01N21/64C12Q1/02
CPCG01N21/6428G01N33/5047
Inventor 韩莉
Owner 郑州鲲鹏健康科技有限公司
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