Method for detecting lymphopoiesis by utilizing medicament cytochalasin B

A technology of lymphocyte proliferation and cytochalasin, which is applied in the field of biological detection, can solve the problems of limited laboratory research, not suitable for clinical use, and expensive CD molecule antibodies of flow cytometry, and achieves low experimental cost, low equipment requirements, and high cost. Simple to use effects

Inactive Publication Date: 2014-09-03
FUDAN UNIV
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Problems solved by technology

This method can detect the reactivity of different subgroups of cells to polyclonal or specific stimuli, and has the advantages of high automation and high throughput. It is currently the most common detection method for lymphocyte proliferation, but flow cytometry and various CD molecular antibody is expensive
[0007] Due to the defects of the above conventional methods themselves, a series of problems will be encountered in practical application; in addition, due to their complexity and precision, these conventional methods are absolutely not suitable for clinical use and can only be limited to laboratory research

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  • Method for detecting lymphopoiesis by utilizing medicament cytochalasin B

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Embodiment Construction

[0029] (1) Materials

[0030] 1. Phytohemagglutinin (Sigma Company), cytochalasin B (Sigma Company).

[0031] 2. The culture medium is RPMI1640 (PAA), containing 10% calf serum (Sijiqing), and 1% double antibody (Geno).

[0032] 3. Heparin anticoagulation tube.

[0033] 4. Normal saline, 0.87% NH 4 Cl solution, Giemsa staining solution.

[0034] (2) Operation method

[0035] 1. Sterile blood collection, 0.5ml each, divided into 2Gy irradiation group (proliferation inhibition group) and non-irradiation group, respectively injected into 4.5ml RPMI1640 culture medium containing PHA, the final concentration of PHA was 50μg / ml, mixed well, placed in Incubate in a 37°C incubator for 2 days.

[0036] 2. On the third day, add cytochalasin B to the culture medium with a final concentration of 3 μg / ml, and continue to culture the cells for 24 hours.

[0037] 3. Dissolve red blood cells: Discard the supernatant on the fourth day and add 0.87% NH prewarmed at 37°C 4 Cl solution 3...

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Abstract

The invention belongs to the technical field of biological detection, and particularly relate to a method for detecting lymphopoiesis by utilizing a medicament cytochalasin B. The principle of the invention is that the medicament cytochalasin B can retard cell division and cannot retard nuclear division, so that cells which are subjected to breeder reaction are treated by the cytochalasin B to form visual binucleated cells; peripheral blood lymphocyte can enable corresponding lymphocyte to be cloned and multiplicated under the stimulation of phytohemagglutinin, and ionizing radiation can inhibit multiplication of lymphocyte; human peripheral blood can be irradiated by a 2Gygamma ray, and the cytochalasin B is added to treat after the stimulation by PHA (phytohemagglutinin), and difference of binuclear rate of radiated and non-radiated lymphocyte is detected; the lymphocyte stimulated by PHA forms binucleated cells, and the binuclear rate of radiated lymphocyte decreases obviously, which shows that the multiplication level of lymphocyte can be reflected well by detecting the binuclear rate of lymphocyte by utilizing the cytochalasin B. The method provided by the invention is visual in detecting indexes, is simple, rapid and economical in operation, and is accurate in result.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and in particular relates to a method for detecting lymphocyte proliferation. Background technique [0002] Lymphocyte proliferation and differentiation is an important stage in the body's immune response process. Therefore, detecting the level of lymphocyte proliferation is a common method for cellular immunity research and clinical immune function detection. It is a must for medical students, especially laboratory students. experimental project. Clinically, it is mainly used to detect the effects of various drugs on immune function and to diagnose diseases that may involve cellular immunity, such as tumors, delayed allergy, and cellular immune deficiency. [0003] Many complex biochemical reactions occur during the proliferation and activation of lymphocytes, including early membrane-associated phenomena such as increased phospholipid synthesis, increased permeability of divalent ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/02
CPCG01N33/5047
Inventor 袁德晓邵春林张江虹潘燕白杨
Owner FUDAN UNIV
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