#immunochromatography

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Wes fully automated protein expression quantitative analysis system

Colloidal gold is a commonly used labeling technique. It is a new type of immunolabeling technique that uses colloidal gold as a tracer to be applied to antigen and antibody. It has its unique advantages. It has been widely used in various biological researches in recent years. Almost all immunoblotting techniques used in the clinic use its markers. At the same time, it may be used in flow cytometry, electron microscopy, immunology, molecular biology and even biochips.

Faulk and Taytor introduced colloidal gold into immunochemistry in 1971. Since then, immunocolloidal gold technology has been increasingly used in various fields of biomedicine as a new immunological method. The current applications in medical tests are mainly immunochromatogra-phy and Dot-immuogold filtration assay DIGFA, which are used to detect HBsAg, HCG and anti-double-stranded DNA antibodies, etc., with simple , Fast, accurate and pollution-free.

1. The basic principles of immune colloidal gold technology

Colloidal gold is made of chloroauric acid (HAuCl4) under the action of reducing agents such as white phosphorus, ascorbic acid, sodium citrate, tannic acid, etc., which can be polymerized into gold particles of a certain size, and become a stable colloidal state due to static electricity. It forms a negatively charged hydrophobic glue solution, which becomes a stable colloidal state due to static electricity, so it is called colloidal gold. Colloidal gold is negatively charged in a weak alkaline environment and can form a firm bond with the positively charged groups of protein molecules. Since this bond is electrostatic, it does not affect the biological properties of the protein.

In addition to binding to proteins, colloidal gold can also bind to many other biological macromolecules, such as SPA, PHA, ConA, etc. According to some physical properties of colloidal gold, such as high electron density, particle size, shape and color response, coupled with the immune and biological properties of the conjugate, colloidal gold is widely used in immunology, histology, pathology and cells Biology and other fields.

Colloidal gold labeling is essentially a coating process in which polymers such as proteins are adsorbed onto the surface of colloidal gold particles. The adsorption mechanism may be the negative charge on the surface of the colloidal gold particles, which forms a firm bond with the positively charged groups of the protein due to electrostatic adsorption. The reduction method can be used to conveniently prepare colloidal gold particles of various sizes, that is, different colors, from chloroauric acid. This spherical particle has a strong adsorption function for protein, and can bind non-covalently with Staphylococcus A protein, immunoglobulin, toxin, glycoprotein, enzyme, antibiotic, hormone, bovine serum albumin polypeptide conjugate, etc. Therefore, it becomes a very useful tool in basic research and clinical experiments.

Immunogold labelling technology (Immunogold labelling techique) mainly utilizes the high electron density of gold particles. At the junction of gold-labeled protein, dark brown particles can be seen under the microscope. When these labels aggregate in large amounts at the corresponding ligands, Red or pink spots are visible to the naked eye, so it is used in qualitative or semi-quantitative rapid immunoassay methods. This reaction can also be amplified by the deposition of silver particles, which is called immunogold-silver staining.

2. Commonly used immune colloidal gold detection technology

(1) Immunocolloidal gold microscope staining method

Cell suspension smears or tissue sections can be stained with colloidal gold-labeled antibodies, or on the basis of colloidal gold labeling, with silver developing solution to enhance the labeling, so that the reduced silver atoms are deposited on the surface of the labeled gold particles. Can significantly enhance the sensitivity of colloidal gold labeling.

Immunogold electron microscopy staining

Colloidal gold-labeled antibodies or anti-antibodies can be combined with negatively stained virus samples or tissue ultrathin sections, and then negatively stained. It can be used for the observation of virus morphology and virus detection.

(2) Dot immunogold percolation method

Use a microporous filter membrane (such as a membrane) as a carrier, first spot the antigen or antibody on the membrane, add the sample to be tested after sealing, and then use the colloidal gold-labeled antibody to detect the corresponding antigen or antibody.

(3) Colloidal gold immunochromatography

IgM mainly exists in the blood of organisms and plays a major role in preventing bacteremia. Therefore, the determination of IgM antibodies has high clinical diagnostic value for certain infectious diseases. IgM accounts for 5% to 10% of the total serum immunoglobulin, and the serum concentration is about 1 mg/ml. Monomer IgM is expressed on the surface of B cells in a membrane-bound form and constitutes a B cell antigen receptor. Igm is the earliest antibody that appears in the initial humoral immune response and is the forerunner of the body's anti-infection. IgM detected in the serum indicates the recent occurrence of infection and can be used for early diagnosis of infection.

Professor Wang Chuanxin of the Second Hospital of Shandong University, together with Shandong Normal University and Weifang Kanghua Biotechnology Co., Ltd. established a joint R&D team to develop a new coronavirus (2019-nCoV) IgM antibody detection kit (colloidal gold method and immunochromatographic method). Results will be available within 3 minutes at the fastest.

Colloidal Gold Method

Under the action of reducing agents such as white phosphorus, ascorbic acid, sodium citrate, tannic acid, chloroauric acid (HAuCl4) can polymerize into gold particles of a certain size, and become a stable colloidal state due to static electricity, forming a negative charge Hydrophobic glue solution of phosphate becomes a stable colloidal state due to static electricity, so it is called colloidal gold. Colloidal gold is also an ideal immunological marker in immunoelectron microscopy.

The basic principle of immune colloidal gold technology:

Colloidal gold is negatively charged in a weak alkaline environment and can form a firm bond with the positively charged groups of protein molecules. Since this bond is electrostatic, it does not affect the biological properties of the protein. In addition to binding to proteins, colloidal gold can also bind to many other biological macromolecules, such as SPA, PHA, ConA, etc. Coupled with the immune and biological properties of the conjugate, colloidal gold is widely used in immunology, histology, pathology and cell biology. The reduction method can be used to conveniently prepare colloidal gold particles of various sizes, that is, different colors, from chloroauric acid. This spherical particle has a strong adsorption function for protein, and can bind non-covalently with Staphylococcus A protein, immunoglobulin, toxin, glycoprotein, enzyme, antibiotic, hormone, bovine serum albumin polypeptide conjugate, etc. Therefore, it becomes a very useful tool in basic research and clinical experiments.

Commonly used immune colloidal gold detection technology:

1. Immunocolloidal gold microscope staining method

2. Immunogold electron microscope staining method

3. Use a microporous filter membrane (such as a membrane) as a carrier, first spot the antigen or antibody on the membrane, add the sample to be tested after sealing, and then use the colloidal gold-labeled antibody to detect the corresponding antigen or antibody.

4. Colloidal gold immunochromatography

Immunochromatography