Cytokine is a general term for biologically active small molecule protein substances secreted by cells. In the process of immune response, cytokines play an important role in physiological and pathological processes such as immune regulation, inflammatory response, and tumor metastasis. The detection of cytokines is not only a means of basic immune research, but also has important value in clinical disease diagnosis, disease course observation, efficacy judgment and cytokine therapy monitoring. However, due to the small content of cytokines in the body, the detection of cytokines has brought difficulties. The detection of cytokines has not been widely carried out in clinical diagnosis. It is known that the currently used cytokine detection methods are imperfect and different. The results obtained by the detection method are quite different, which brings certain difficulties to clinical diagnosis and treatment. Therefore, it is necessary to understand the characteristics and influencing factors of various detection methods. At present, there are mainly the following types of cytokine biological activity detection and concentration determination methods. 1. Biological testing Biological testing, also known as biological activity testing, is a test designed based on the specific biological activity of cytokines. Since various cytokines have different activities, such as IL-2 promotes lymphocyte proliferation, TNF kills tumor cells, and CSF stimulates hematopoietic colony formation (Shanghai Chuangsai Technology provides rHuG-CSF recombinant human granulocyte colony-stimulating factor, commodity number: C22-102-02-2μg, price 510 yuan), IFN protects cells from virus attack (Shanghai Chuangsai Technology provides rHuIFN-ω recombinant human interferon ω, commodity number: C22-106-07-5μg, price 510 yuan) Therefore, by selecting the unique biological activity of a certain cytokine, it can be detected. Biological activity detection methods can be further divided into the following categories: Cell proliferation method Many cytokines have cell growth factor activity, particularly interleukins, such as IL-2 stimulates t cell growth, IL-3 stimulates mast cell growth, IL-6 stimulates plasma cell growth, and the like. Using this property, cells that respond to specific cytokines have been screened, and cell lines that depend on only one factor, that is, cell-dependent strains (referred to as dependent strains) have been established. These dependent strains are not normally viable and can only proliferate after the addition of specific factors. For example, the IL-2 dependent strain ctll-2 dies quickly in the medium without IL-2, and after IL-2 is added, it can be cultured in the right body for a long time. In a certain concentration range, cell proliferation is directly proportional to the amount of IL-2, so the content of IL-2 is identified by measuring cell proliferation (eg, using 3h-tdr incorporation method, MTT method, etc.). In addition to dependent strains, there are also short-term cultured cells, such as thymocytes, bone marrow cells, and lymphoblasts stimulated by mitogens, which can be used as target cells to measure certain cytokine activity. Shanghai Chuangsai Technology provides rHuIL-2 recombinant human interleukin-2, product number: C22-101-02-10μg, price 510 yuan. 2. Target cell killing method It is a detection method designed according to the ability of certain cytokines (such as TNF) to kill target cells in vitro. Generally, the target cells are selected from tumor cell lines that have been passaged in vitro for a long time, and the killing rate of the cells is determined by an isotope release method or a dye staining method. Shanghai Chuangsai Technology provides rHuTNF-α/TNFSF2, recombinant human tumor necrosis factor-α/tnfsf2, commercial number: C22-103-01-10μg, price 510 yuan. 3. Cytokine induced product analysis Certain cytokines can stimulate specific cells to produce biologically active substances, such as IL-2, IL-3 induces synthesis of amines in bone marrow cells, and IL-6 induces synthesis of α1-antichymotrypsin by hepatocytes. The activity of the cytokine can be reflected by measuring the corresponding product induced. 4. Cytopathic inhibition The virus can cause damage to the target cells, and interferon can inhibit the cytopathic effect caused by the virus, so the cytopathic inhibition method can be used to detect such factors. Second, immunological detection Cytokines are all proteins or polypeptides and have strong antigenicity. With the advent of recombinant cytokines, it is convenient to obtain specific anti-serum or monoclonal antibodies of cytokines, so that the cytokines can be quantitatively detected by immunological techniques using the characteristics of antigen-antibody-specific reactions. Despite the wide variety of cytokines, similar techniques can be used to work with specific antibodies (including polyclonal or monoclonal antibodies) for a certain factor. Common methods include ELISA, RIA, and immunoblotting. Currently, almost all common cytokine detection kits are commercially available. In addition, enzyme-labeled or fluorescently labeled anti-cytokine monoclonal antibodies can be used to detect the synthesis and distribution of factors in cells in situ, such as the recent development of intracellular staining and enzyme-linked immunospot (ELISPOT) technology. . The immunological assay can directly determine the specific cytokine content (in ng/ml) of the sample, which is convenient for large-scale detection of cytokine levels in serum of clinical patients. This method only measures the antigenicity of cytokines, and is not necessarily parallel to the activity of the factor. Therefore, in order to understand the biological effects of cytokines, it is necessary to combine biological detection methods. Shanghai Chuangsai Technology provides human interleukin-12Elisa kit, Human Interleukin-12, IL-12 ELISA Kit, product number: LH-E10217HU-96T, price 1700 yuan. Third, molecular biology methods This is a technique for detecting the expression of specific cytokine genes using gene probes for cytokines. At present, all the genes of the recognized cytokines have been cloned, so that it is easier to obtain a cDNA probe of a certain cytokine or artificially synthesize an oligonucleotide probe based on a known nucleotide sequence. There are various methods for detecting cytokine mRNA expression by using gene probes, such as dot blot hybridization, Northern blot, reverse transcription PCR, and in situ hybridization of cells or tissues. The key to the experiment is to prepare high quality nucleic acid probes and obtain acceptable analytes (extracted mRNA samples or cell/tissue specimens). A nucleic acid probe refers to a DNA fragment or single-stranded DNA or RNA which is labeled with a radioisotope or other label (such as biotin, digoxin, etc.) and is complementary to the gene of interest. According to their sources, they can be classified into cDNA probes, oligonucleotide probes, genomic gene probes, and DNA probes. Among them, cDNA probes and synthetic oligonucleotide probes are commonly used for dot blot hybridization and Northern blot, and RNA probes are more suitable for in situ hybridization because of their good penetrability. The application of nucleic acid probe technology has been programmed. The main examples of cDNA probes include: 1 extraction of plasmid DNA; 2 separation of target DNA fragments; 3 target DNA fragment labeling; 4 extraction of mrna sample to be tested; Hybridization for the sample to be tested; 6 autoradiography or color analysis. The method of detecting specific mRNA by RT-PCR which has appeared in recent years is also widely used in the field of cytokine research. The method has the advantages of sensitivity and rapidity, and even the specific mRNA can be detected from 1 to 10 cells. The above three methods each have advantages and disadvantages, and can complement each other. In practical applications, they can be selected according to their respective experimental purposes and laboratory conditions. The biological detection method is sensitive and can directly measure biological functions. It is the most reliable method and is suitable for various experimental purposes. It is the most commonly used technology in scientific research departments, but it requires long-term cultivation of dependent cell lines, and the detection takes a long time. It is complicated and has many influencing factors, so it is not easy to master. The immunological assay is relatively simple, rapid, and reproducible, but the measured amount represents only the amount of the corresponding cytokine and does not represent activity, and the sensitivity is also lower than the biological activity assay (about 10 to 100 times lower). Molecular biology can only detect the expression of genes, and can not directly provide information on the concentration and activity of related factors, mainly used for mechanism discussion. In the detection of cytokines, it must be considered that the role of cytokines has a network characteristic. It is necessary to clarify the cytokine components determined by the detection method, and consider the levels of inhibitors and soluble receptors, and combine them in various combinations. It is possible to get more reliable results. Shanghai Chuangsai Technology has excellent performance, interleukin cytokines, fetal bovine serum, electrophoresis equipment scientific instruments, raw material drug standards, chemical reagents, cell culture consumables, Shanghai Chuangsai, mass products special promotions, welcome to inquire! 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