Application of immunomagnetic beads in the field of biomedicine

Immunomagnetic bead technology is a technical method that appeared in the 1980s. Based on immunology, it has penetrated into various fields such as pathology, physiology, pharmacology, microbiology, biochemistry, and molecular genetics. It has become more and more widely used in immunoassays, cell separation, purification of biological macromolecules, and molecular biology.

Structure of immunomagnetic beads

Magnetic beads are composed of core metal particles (Fe2O3, Fe3O4), a polymer material (such as polystyrene, polyvinyl chloride) wrapped in the core outer layer, and the outermost functional ligand (such as -NH2, -COOH, -OH, -CHO) composition.

Application of immunomagnetic beads in the field of biomedicine

Nucleic acid purification magnetic beads NGS screening

1. Cell sorting

Immunomagnetic bead cell sorting can separate very high purity cells from complex cell mixtures within a few minutes. When using nano-scale magnetic beads for cell sorting, the size of the magnetic beads and its composition make it biodegradable without activating the cells or affecting the function and vitality of the cells, and the physiological functions of the cells are also unchanged. Magnetically labeled cells can be used immediately for analysis and subsequent experiments.

Cell sorting can be divided into

a) Positive sorting (positive sorting): The cells bound by magnetic beads are the cells to be separated, which is suitable for flow analysis and cell-based analysis.

b) Negative sorting (negative sorting): The magnetic beads bind to unwanted cells, and the cells free in the supernatant are the desired cells.

Positive selection (left) and negative selection (right) are as follows:

1. Important indicators for magnetic separation of cells

Purity and yield depend on the specificity of the monoclonal antibody connected to the magnetic beads and the size (magnetic) of the magnetic beads. However, the yield of magnetic beads that are too small is not high, and magnetic beads that are too large will affect cell viability and cannot directly Upstream.

2. Protein/antibody separation and purification

The application of antibody (protein)-coated immunomagnetic bead purification technology does not require complicated chromatography equipment, and has no limitation on the clarity of the sample. It only requires a simple magnetic adsorption step to easily separate the monoclonal antibody from the monoclonal antibody expression product. Effectively solve the shortcomings of traditional chromatography technology.

3. Nucleic acid separation and purification

The binding of nucleic acid to magnetic beads mainly relies on electrostatic, hydrophobic and hydrogen bonding. The DNA/RNA in the cell or tissue is released under the action of the lysis solution. At this time, the surface-modified superparamagnetic silica nanomagnetic beads “specifically bind” with nucleic acid to form a “nucleic acid-magnetic bead complex”. Then under the action of an external magnetic field, the complex is separated.

It can be widely used in genome research in molecular biology, molecular evolution research, genetic disease research in medicine, mutation gene detection, tumor screening, HPV detection, HLA typing, transplantation matching, etc., forensic biological samples The detection of blood spots, fine spots, hair, cigarette butts and other on-site evidence, and judicial paternity testing, blood relationship identification, etc. provide evidence, archeology, biological experiments in universities, middle schools, and many other fields.

Amino Magnetic Beads

Magnetic bead method to extract nucleic acid:

Traditional nucleic acid extraction method Magnetic Bead Extraction of Nucleic Acid
Not technically difficult High quality, high yield, high throughput
Manual extraction Realize process automation
The operation is cumbersome, time-consuming and laborious Eliminates complicated manual extraction procedures
Not suitable for nucleic acid extraction of a large number of samples Reduce the damage of phenols, chloroform and other organic reagents to operators
Not suitable for clinical molecular diagnosis Greatly meet today’s market requirements for nucleic acid extraction efficiency

4. Immunoassay

Because of its small particle size and large specific surface area, immunomagnetic beads can capture more analytes, and directly perform enzyme color, fluorescence or isotope display on their surface, thus establishing a series of fast detection speed, high specificity, and sensitivity. High and reproducible immunoassay method.

5. Other applications

Under a high gradient magnetic field, the immunomagnetic bead method is used to separate B and T lymphocytes in abdominal blood or veins. The separated lymphocytes are then widely used in the rapid selection of clinical organ transplantation donors and recipients. Perform HLA-I type II antigen typing.

Magnetic bead technology, as a product developed by the convergence of interdisciplinary subjects, plays a huge role in biological and medical laboratory testing. With the rapid development of second-generation sequencing, the consumables involved in second-generation sequencing have also ushered in its spring.

Whether it is magnetic beads used to extract nucleic acids or immunomagnetic beads used in cell sorting and immunoassays, they are still a subdivision of the industry in the field of molecular biology, and the technology is still in continuous progress and development. As more and more companies join, the prices of products developed by them will also become cheaper.

How to perform nucleic acid extraction

The common difficulty in nucleic acid extraction lies in the difficulty of extracting high-quality RNA from some plant tissues, which is related to the rich ingredients in these plant tissues, such as phenolic compounds, polysaccharides, and some unidentified secondary metabolites. In addition, there are tissues that are rich in higher activity. In an intact cell, these substances are separated from nucleic acids. Once the cell is broken, these substances will interact with RNA.

Nucleic acid extraction method:

1. Quickly inactivate endogenous RNase to prevent RNA degradation. The following 3 methods can effectively inactivate endogenous RNase:

1) Harvest the sample with a cell lysate containing chaotropic (such as guanidine salt) and homogenize immediately.

2) Instantly freeze the sample with liquid nitrogen. It is worth noting that: the tissue pieces must be small enough to be frozen the moment they are immersed in liquid nitrogen to ensure instant inactivation of RNase.

3) Immediately place the sample in the liquid nitrogen-free RNA sample storage solution. It is an aqueous, non-toxic collection reagent that can immediately stabilize and protect RNA in intact, unfrozen tissue and cell samples. The key point is that the tissue sample slice must be thin enough (<0.5 cm) so that it can quickly penetrate into the tissue block before the RNase destroys the RNA.

2. Use the correct cell or tissue storage conditions

After the sample is frozen instantaneously with liquid nitrogen, it should be stored at -80°C and must not be thawed. Even a short thawing before homogenization in a lysate containing guanidine salt will result in degradation and loss of RNA. Instantly frozen tissues should be ground into powder under ultra-low temperature, and then placed in the lysis solution for homogenization.

Manual nucleic acid extraction kit

3. Choose a good RNA isolation method

The numerous RNA isolation methods available may be difficult to choose. The current simple and safe method is column separation. For example, it is loved by everyone because of its simple operation, short time and high purity. It does not require DNase to digest DNA, which saves time and avoids RNA degradation, thereby increasing the yield of nucleic acid extraction; Relative to non-column separation, it removes proteins and other impurities cleanly, improves purity, and is very suitable for cells, tissues, and general plants.

Plant nucleic acid extraction is more difficult to choose. Plant RNA extraction is affected by the content of phenols, polysaccharides, protein impurities, and secondary metabolites. You can choose the polysaccharide and polyphenol plant total RNA extraction kit (narcissus, pepper, carrot, corn, lily, wheat, Tomatoes, cauliflower, rapeseed, etc.) and general plant RNA extraction kits (suitable for the extraction of most plants, including: apple, grape, strawberry, banana, longan, lychee, lawn plant, pine, cedar, white birch, purple pine cone, Coleus, poinsettia, oleander, Ficus benjamina, violet, rose, geranium, morning glory, etc.); it is worth mentioning that blood (including serum, plasma, cerebrospinal fluid, other liquids) nucleic acid extraction TRIZOL and red blood cells The lysis method is not effective. The red blood cell lysate does not contain RNase inhibitors. In this process, RNA is easily degraded. It is recommended to use a blood total RNA extraction kit.

What are the precautions for the nucleic acid extraction system when doing separation experiments?

The nucleic acid extraction system is a high-tech product that uses the universal magnetic bead method to extract nucleic acids. It has the advantages of high automation, fast extraction speed, stable results, and easy operation. In almost every laboratory, the separation and purification work related to biomolecules is very important.

However, it is quite difficult to purify multiple samples. It is not only necessary to select a suitable purification technology, but also the workload is particularly large, which is difficult to meet the current rapid development of high-throughput sample extraction and purification needs.

The magnetic bead method of nucleic acid extraction system is very suitable for genomics research. Regardless of whether the source of the extracted sample is microorganisms, animals, plants or viruses, combined with a whole blood genomic DNA extraction kit specially optimized for magnetic bead extraction and purification systems, a white blood cell layer whole blood genome extraction kit, and animal tissue/cell genomic DNA extraction reagents The box can quickly purify DNA or RNA in sufficient quantity and purity.

HERO96 magnetic bead method nucleic acid extraction system

 

What should we pay attention to when using a nucleic acid purifier for separation experiments?

1. Ensure the integrity of the primary structure of nucleic acid.

2. Remove the pollution of other biological macromolecules (protein, carbohydrate, lipid molecules).

3. Remove the contamination of other nucleic acid molecules (extract DNA and remove RNA contamination).

4. Remove organic solvents and high-concentration metal ions that inhibit enzymes.

5. Simplify the operation steps and shorten the extraction process.

HERO 32 Magnetic Bead Method Nucleic Acid Extraction System

6. Prevent the degradation of nucleic acid by physical and chemical factors (shear force, high temperature, strong acid and strong base).

7. Prevent the degradation of nucleic acid by biological factors (nuclease).

The nucleic acid purification instrument has stable measurement results, avoids differences and errors caused by manual operation, and has good temperature and repeatability. The purifier can optimize the purification scheme according to the reagents and cooperate with the precise incubation time to achieve higher extraction efficiency.The extracted DNA/RNA has high purity and can be directly used for PCR and RT-PCR.

This product has a powerful program editing function; it can define your application flexibly and efficiently to meet the requirements of different reagents. The lysis and elution temperature can be customized according to the needs. Built-in engineering computer, no need to connect to a personal computer; stand-alone operation saves more space and energy, and provides a highly stable automatic control system.

Basic magnetic beads common questions and answers

Magnetic beads is a newly developed immunological technology in recent years. It combines the unique advantages of curing reagents with the high specificity of immunological reaction. Based on immunology, it penetrates into pathology, physiology, pharmacology, microorganisms, biochemistry and molecules. Genetics and other fields, and have been more and more widely used in immunological detection, cell separation, biological macromolecule purification and molecular biology. Common types of magnetic beads include hydroxyl magnetic beads, amino magnetic beads and carboxyl magnetic beads. Different types of magnetic beads have different application problems. Next, the editor of Aisen will share with you some common questions about basic magnetic beads and the answers to these questions.

Frequently Asked Questions about Hydroxyl Magnetic Beads

1. What is the dispersion of each type of magnetic beads?

70301 Mag OH-500 has uniform particle size and monodisperse;

70302 Mag OH-1000 amorphous, non-uniform particle size, average particle size 1 micron, polydisperse;

70802 Magrose OH spherical, particle size 30-150 microns, polydisperse.

2. How much is the nucleic acid binding capacity?

The binding force is greater than 20μg DNA/mg magnetic beads.

3. Application areas of Magrose OH

The surface modification of Magrose OH magnetic beads is used in the field of protein purification.

4. Is there any instructions for using hydroxy magnetic beads for nucleic acid extraction?

Hydroxyl magnetic beads need different buffers for nucleic acid extraction according to different samples. Therefore, there is no relevant instructions, and customers need to choose the buffer by themselves. If you need a matching kit, you can choose the relevant nucleic acid extraction kit.

Frequently Asked Questions about Amino Magnetic Beads

1. 15% glutaraldehyde solution preparation method and matters needing attention

Use PBS buffer for glutaraldehyde, pay attention to the current use.

2. How to deal with biological ligands such as IgG

IgG is prepared with PBS buffer. If there are other small molecules containing carboxyl or amino groups, remove them as much as possible to avoid competing reactions with magnetic beads.

Common problems of carboxyl magnetic beads

1. What is the amount of antibody coupling?

About 2.5-3μg/mg magnetic beads.

2. After carboxyl magnetic beads are coupled to IgG, which magnetic bead storage solution is more suitable?

The preservation solution can be PBST or tris-HCl.

The above is the introduction to the common questions and answers of basic magnetic beads. Different magnetic beads will always encounter various problems in the application process. It is necessary to understand the simple application methods and techniques of various magnetic beads to ensure that the magnetic beads are normal. use. The magnetic beads produced by Aisen Biotechnology are diverse in variety and excellent in performance. Customers in need are welcome to call for consultation.

How accurate is the nucleic acid detection?

Commonly used infectious disease laboratory tests include microbial culture, virus isolation, nucleic acid testing, and antibody testing. Nucleic acid testing is also a method to detect viruses, which can detect infections earlier. Nucleic acid tests have been carried out for hepatitis B, hepatitis C, and AIDS in clinical practice, which can detect the presence of viruses in the blood before antibodies are produced. For some respiratory diseases, nucleic acid testing of throat swabs or respiratory secretions can be used to determine whether they are infected, and whether they are contagious. Seasonal influenza, avian influenza, coronavirus infections, etc., can be easily judged whether there are viral components in the upper respiratory tract through throat swab testing, so as to determine whether the tested person is an infected person. So, how accurate is the nucleic acid detection?

At present, there are no exact large-sample statistics to prove the accuracy of nucleic acid detection, but there are many factors that affect the accuracy, such as the quality of the detection reagents, the time of infection of the tested person (too early or too late will reduce the accuracy), specimens The cooperation of the examinee when taking the material (poor cooperation may cause the specimen not to be taken in place) and the operational proficiency of the examiner, etc.

Criminal Case Detection

If all aspects of nucleic acid detection are accurate, the accuracy of nucleic acid detection can reach more than 95% under ideal conditions. But due to various reasons, false negative results will occur. The so-called false negative means that the testee is indeed an infected person, but the nucleic acid test result is negative, which is a false negative. With the improvement of technology, the accuracy rate of various medical institutions is improving rapidly, and the false negative rate is continuously decreasing.

In addition, for people who have had close contact with confirmed or suspected cases, two nucleic acid tests are usually required to exclude them. For people who have been to high-risk areas of the new coronavirus, if they have fever and respiratory symptoms, a negative nucleic acid test may have a false negative, so two nucleic acid tests are needed to rule out. In addition, if the doctor judges that a nucleic acid test is negative and still cannot be ruled out, the nucleic acid test needs to be tested again. Sometimes the doctor will also recommend chest CT, blood tests, and serum antibody tests for the new coronavirus to determine whether it is infected.

Advantages and uses of common magnetic beads

High-quality magnetic beads have the characteristics of uniform size distribution, high surface loading, extremely short magnetic response time, high dispersion stability, etc., which can quickly and efficiently separate the test object from the sample, greatly improving the detection efficiency. In addition, there are many types of magnetic beads, and different magnetic beads have different advantages and different uses. Next, the editor of Aisen will share with you the relevant knowledge about the advantages and uses of common magnetic beads.

Advantages and uses of carboxyl magnetic beads

1. Advantages

(1) It can be coupled with biomolecules such as protein, nucleic acid, streptavidin, polypeptide, enzyme, etc. through the activation of EDCNHS;

(2) 1μm in size, uniform size, granular roughness on the surface, high specific surface area, extremely high carboxyl loading capacity, and low non-specific adsorption capacity;

(3) It has superparamagnetism, fast magnetic response speed, good monodispersity, and can ensure the uniformity of the reaction and the consistency of detection;

(4) It has a highly hydrophilic surface and good biocompatibility;

(5) Excellent suspension, slow semi-sedimentation speed, suitable for automatic operation.

2. Purpose

Chemiluminescence, gene capture, protein purification, cell sorting.

Advantages and uses of streptavidin magnetic beads

1. Advantages

(1) It can be used for the coupling of biotinylated proteins, nucleic acids, polysaccharides, lipids, enzymes and other biomolecules immediately;

(2) 1μm in size, uniform size, granular roughness on the surface, high specific surface area, extremely high streptavidin loading and biotinylated protein/nucleic acid coupling rate;

(3) Surface modification of hydrophilic polymer molecules and covalent coupling of multi-layer streptavidin. Streptavidin is densely adsorbed on the surface with few exposed sites on the surface, which greatly reduces non-specific adsorption ;

(4) It has good suspension and is stable within pH (3.0-9.0), and can be used for automated high-throughput experimental operations.

2. Purpose

Chemiluminescence, protein and nucleic acid separation.

Advantages and uses of amino magnetic beads

1. Advantages

(1) It can be coupled with biological molecules such as proteins, nucleic acids, polysaccharides, lipids, enzymes, etc. through a variety of coupling methods;

(2) 1μm in size, uniform size, granular roughness on the surface, high specific surface area, and extremely high amino loading;

(3) It has superparamagnetism, fast magnetic response speed, good monodispersity, and can ensure the uniformity of the reaction and the consistency of detection;

(4) It has a highly hydrophilic surface and good biocompatibility;

(5) Excellent suspension, slow semi-sedimentation speed, suitable for automatic operation.

2. Purpose

Chemiluminescence, cell, virus, nucleic acid, antibody separation and purification.

The above is the introduction of the advantages and uses of common magnetic beads. Different magnetic beads have different advantages and performances, and their uses are also different. The suitable magnetic beads should be selected according to their performance characteristics. The magnetic beads produced by Aisen Biotechnology are rich in varieties and excellent in performance. Customers in need are welcome to call for consultation.

What is the principle of magnetic bead method for nucleic acid extraction

With the rapid development of genetic diagnosis, genetically modified food detection, personalized medicine, etc., the current nucleic acid extraction technology can no longer meet the needs of today’s biotechnology, and there is an urgent need for a high-throughput and automated nucleic acid extraction method. In this context, the magnetic bead method for nucleic acid extraction came into being.

The use of magnetic beads for DNA extraction is one of the areas where their biological value is maximized. The main types of microspheres used include silanol magnetic beads and oligo magnetic beads. Magnetic beads with surface groups can specifically and reversibly bind to nucleic acids released in the test sample. At the same time, the magnetic response capability of the magnetic beads is used to carry out directional movement and enrichment under the action of an external magnetic field, so as to realize the separation and purification of nucleic acid. The main advantages of the magnetic bead method for nucleic acid extraction include simple and automated operation, large-scale operation, and short time.

1. What is a magnetic bead

Magnetic beads are a kind of special nano-micron materials. Their size is usually between a few tenths of a few microns to a few microns, which is only one tenth to one tenth of the diameter of a human hair, which cannot be observed by the naked eye. Single magnetic bead. They are superparamagnetic, they can move quickly to one side in a magnetic field, gather together, and can quickly return to a dispersed state after removing the magnetic field.

The surface of the magnetic beads used for nucleic acid extraction has specific properties. Under certain conditions, the viral nucleic acid can be adsorbed on the surface, and under other conditions, the viral nucleic acid can be released from the surface for subsequent detection steps.

2.the principle of magnetic bead method of nucleic acid extraction

The binding of nucleic acid to magnetic beads mainly relies on electrostatic, hydrophobic and hydrogen bonding. The DNA/RNA in the cell or tissue is released under the action of the lysis solution. At this time, the surface-modified superparamagnetic silica nanomagnetic beads “specifically bind” with nucleic acid to form a “nucleic acid-magnetic bead complex”. Then under the action of an external magnetic field, the complex is separated. After the eluate is washed to remove the non-specifically adsorbed magazines, desalted, and purified, the nucleic acid substance to be extracted is obtained.

According to the same principle as the silica gel membrane spin column, the surface of superparamagnetic nanoparticles is modified and modified by nanotechnology to prepare superparamagnetic silica nanomagnetic beads. The magnetic beads can specifically recognize and efficiently bind with nucleic acid molecules on the microscopic interface. Using the superparamagnetism of silica nanospheres, under the action of Chaotropic salts (guanidine hydrochloride, guanidine isothiocyanate, etc.) and an external magnetic field, DNA and DNA can be separated from samples such as blood, animal tissues, food, and pathogenic microorganisms. RNA can be used in clinical disease diagnosis, blood transfusion safety, forensic identification, environmental microbiological testing, food safety testing, molecular biology research and other fields.

In fact, since testing institutions need to process a large number of samples every day, most of the nucleic acid extraction steps are carried out by means of an automatic nucleic acid extraction instrument with a magnetic bead method nucleic acid extraction kit.

What are the factors affecting nucleic acid extraction by magnetic beads?

In the past two years, as pathogen detection has become more popular, pathogen nucleic acid detection has gradually entered people’s field of vision and has become the gold standard for pathogen detection. Faced with large quantities of samples, testing results need to be issued in a short period of time, and people’s needs for fast and efficient testing methods are becoming more and more urgent. As an important step of nucleic acid detection, nucleic acid extraction also faces the requirements of throughput and high efficiency. Automatic nucleic acid extraction (magnetic bead method nucleic acid extraction) has been quickly promoted and applied because of its fast, efficient, easy-to-use, safe and non-toxic characteristics, and has become a standard in laboratories. However, during the extraction process, certain factors will affect the extraction results and require the operator’s attention. The editor of Aisen has compiled the relevant knowledge of the factors affecting nucleic acid extraction by magnetic bead method, and let’s share it with you.

Magnetic bead method nucleic acid extraction consists of lysis, washing and elution steps. The factors that affect these steps will also affect the extraction efficiency of magnetic beads. Below we explain each step one by one.

   Factors affecting the lysis step

  1.Lysis solution composition

For some samples that are difficult to lyse, the lysis ability of the lysate can be enhanced by increasing the concentration of guanidine salt and surfactant. Guanidine salts and surfactants can promote protein denaturation, destroy cell membranes, dissociate nucleic acids and proteins, and release more nucleic acids.

  2.pyrolysis temperature and time

Increasing the temperature and increasing the lysis time can make the cell lysis more thoroughly and get more nucleic acids. You need to explore the appropriate temperature and time; too high temperature or too long time may also lead to nucleic acid fragmentation or reduced extraction efficiency caused by alcohol volatilization.

  3.magnetic beads adsorption capacity

magnetic bead extraction series magnetic beads are generally silanol magnetic beads, and the adsorption capacity of the magnetic beads is affected by the specific surface area of ​​the magnetic beads and the density of the modified group. Generally, magnetic beads with more modified groups and larger specific surface area are selected. Generally, the smaller the particle diameter of the magnetic beads, the larger the specific surface area. But it is not that the smaller the particle size of the magnetic beads, the better, the smaller the particle size of the magnetic beads, the adsorption speed will also slow down, you can choose according to the actual sample situation. The binding buffer is generally combined with isopropanol or ethanol. Generally, the binding capacity can be enhanced by increasing the ratio of alcohol, and the concentration of chaotropic salt should be increased to promote the binding of nucleic acid and magnetic beads.

   Factors affecting washing steps

Washing is very important for the entire extraction step. It is related to the purity of nucleic acid, which directly affects downstream PCR applications. The oscillation amplitude and mixing time of the magnetic bar will affect the washing efficiency.

  1.Magnetic rod oscillation amplitude

If the oscillation amplitude of the magnetic rod is too small, there will be impurities remaining on the magnetic beads, which will affect the ability of the magnetic beads to adsorb nucleic acids; if the oscillation is too violent, part of the nucleic acid will be lost or the nucleic acid will be broken or degraded, and the yield will decrease.

  2.mixing time

If the mixing time is too short, there will be impurities remaining on the magnetic beads, which will affect the ability of the magnetic beads to adsorb nucleic acids; if the mixing time is too long, part of the nucleic acid will be lost or the nucleic acid will be broken or degraded, and the yield will decrease.

COVID-19 Virus Nucleic Acid Extraction Kits & System

   The influence of elution step factors

The elution step directly affects the yield of nucleic acid. The factors affecting the elution efficiency are the drying degree of the magnetic beads and the elution temperature and time.

   1.Dryness degree of magnetic beads

If the magnetic beads are not fully dried, there may be impurities such as alcohol, which will affect the subsequent PCR reaction; if the magnetic beads are too dry, the nucleic acid may dry up on the magnetic beads, resulting in a decrease in elution efficiency.

  2.elution temperature and time

Appropriately increasing the elution temperature and time will help the nucleic acid to be released from the magnetic beads. If the time is too long or the temperature is too high, the nucleic acid may be degraded, and the balance needs to be grasped.

The relevant content about the factors that affect the magnetic bead method of nucleic acid extraction is the above. When performing the magnetic bead method of nucleic acid extraction, ensure that the above factors are reasonable, so that the extraction quality can be better. Aisen Biotechnology has extensive experience in the production of nucleic acid extraction systems and nucleic acid extraction kits. It uses high-quality technology and reliable quality. Customers in need are welcome to call for consultation.

Magnetic bead method of nucleic acid extraction steps and principles

With the continuous development of biomedicine, nucleic acid extraction technology is also being updated. Magnetic bead nucleic acid extraction, as a new type of nucleic acid extraction technology, can extract nucleic acids with high throughput and automation, meeting the needs of today’s biotechnology. In addition, the steps of magnetic bead method for nucleic acid extraction are relatively simple, easy to operate, and are well received. Next, I will share with you the steps and principles of magnetic bead method for nucleic acid extraction.

Magnetic bead method for nucleic acid extraction steps

1. Add 1 mL of deionized water to each bottle of 40mg proteinase K dry powder to a final concentration of 40mg/mL, and add 500 μL of deionized water to each bottle of 20mg proteinase K dry powder to a final concentration of 40mg/mL, and mix it upside down. It can be completely dissolved once, and it can be stored at 4℃ for short-term use. Please keep it at -20℃ for long-term storage. Repeated freezing and thawing should not exceed five times.

2. The tissue sample is ground into fine powder with 10-30 mg of liquid nitrogen, transferred to a 1.5 mL centrifuge tube, resuspended in 200 μL of 1 X PBS buffer, and then proceeded to the next step. The liquid sample goes directly to the next step.

3. Take 500μL of virus lysate VL and add it to a 1.5mL enzyme-free centrifuge tube, then add 25μL of proteinase K and 25μL of viral magnetic beads (shaken well before adding), and then add 200μL of serum/plasma/tissue homogenate sample , Vortex and mix well, 55% C water bath for 20 minutes, invert and mix for 10 seconds every 5 minutes.

4. Take out the enzyme-free centrifuge tube in the water bath, place it on the magnetic stand, and magnetize for 90 seconds to completely adsorb the magnetic beads. Use a pipette to carefully aspirate and discard the supernatant (be careful not to attract the magnetic beads at the bottom of the tube).

5. Remove the centrifuge tube from the magnetic stand, add 700μL of rinsing solution BufferA, mix by pipetting or vortex for 30 seconds to resuspend the magnetic beads, then place the centrifuge tube on the magnetic stand, and magnetize for 90S to make the beads Adsorbed completely, carefully aspirate and discard the supernatant with a pipette.

6. Take 700μL of rinsing solution BufferB, add it to the centrifuge tube, try not to blow off the magnetic beads, and then directly aspirate and discard the supernatant.

7. Add 80μL of elution buffer, remove the centrifuge tube, and bathe in a water bath at 56°C for 5 min. During this period, vortex 3-4 times to elute the nucleic acid from the magnetic beads.

8. Place the centrifuge tube on the magnetic stand and magnetically attract for 60 seconds, transfer the liquid to a new 1.5mL enzyme-free centrifuge tube, and proceed directly to the downstream experiment or store at -20°C.

HERO 32

The principle of magnetic bead method for nucleic acid extraction

Its principle mainly relies on electrostatic interaction, hydrophobic interaction and hydrogen bonding interaction. The DNA/RNA in the cell or tissue is released under the action of the lysate. At this time, the surface-modified superparamagnetic silica nanomagnetic beads “specifically bind” with nucleic acid to form a “nucleic acid-magnetic bead complex”. Then under the action of an external magnetic field, the complex is separated. Then the eluate is used to wash away non-specifically adsorbed impurities, desalt and purify to obtain the nucleic acid substance to be extracted.

The above is the introduction to the steps and principles of the magnetic bead method for nucleic acid extraction. The magnetic beads and nucleic acid extraction system produced by Aisen Biotechnology have excellent performance and reliable quality. Customers in need are welcome to call for consultation.

What is nano magnetic bead technology?

Nanomagnetic beads refer to small magnetic particles whose size is suitable to be measured by nanometers, generally 1-100 nanometers. This kind of magnetic beads has a very special magnetic property called superparamagnetism, that is, it has a strong magnetic field in an external magnetic field. Responsiveness, and after the magnetic field is removed, the magnetic properties of the magnetic particles disappear immediately, that is, there is no remanence, and they are uniformly dispersed in the solution again. Traditional DNA separation technology includes precipitation, centrifugation and other processes. These purification methods have complicated steps, time-consuming, low yield, and contact with toxic reagents. It is difficult to realize automatic operation; however, the use of magnetic carrier microsphere separation technology can overcome it well. These shortcomings enable rapid and efficient sample preparation. Therefore, nanomagnetic bead technology is an important direction for the development of DNA purification methods in the future.

What are nano magnetic beads

The so-called nano magnetic beads refer to small magnetic particles whose size is suitable to be measured by nanometers, generally referring to 1-100 nanometers. This kind of magnetic beads has a very special magnetic property called superparamagnetism, that is, it has a strong magnetic field in an external magnetic field. Magnetic responsiveness, and after the magnetic field is removed, the magnetic properties of the magnetic particles disappear immediately, that is, there is no remanence, and they are uniformly dispersed in the solution again.
Scientists favor this feature, which can be used to adsorb a certain component in the liquid, and then magnetically separate the magnetic beads to achieve the purpose of separating the components. Of course, in order to be able to adsorb the desired substance, specific groups must be wrapped on the outside of the particles, such as amino, hydroxyl, carboxyl, sulfhydryl and other functional groups. These groups are specifically combined with the target molecule, and then collected by magnetic force. Magnetic beads can separate the required substances.
The magnetic beads themselves are mostly inorganic materials, the most common material is ferroferric oxide, and the outer covering is basically organic material. According to the position relationship between the magnetic bead itself and the coating material, there are four types of magnetic bead structures, namely, core-shell type, mosaic type, core-shell type, and core-shell type.

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Among them, the first core-shell type has been studied and used the most. Its magnetic material is used as the core, and the polymer material is used as a shell to wrap the outside of the magnetic particles. The inorganic magnetic material at the core endows the entire particle with superparamagnetism. Separation, the high-molecular substance in the shell gives the entire particle high-efficiency adsorption, that is, specificity. In a fully mixed and uniform liquid environment, the polymer shell is combined with the target substance through its functional group, and the movement and collection of the entire particle is controlled by an external magnetic field, and finally the goal of separation of the target substance is achieved.

The use of magnetic beads for DNA extraction is one of the areas where their biological value is maximized. The main types of microspheres used include silanol magnetic beads and oligo magnetic beads. Magnetic beads with surface groups can specifically and reversibly bind to nucleic acids released in the test sample. At the same time, the magnetic response capability of the magnetic beads is used to carry out directional movement and enrichment under the action of an external magnetic field, so as to realize the separation and purification of nucleic acids. The main advantages of the magnetic bead method for nucleic acid extraction include simple and automated operation, large-scale operation, and short time.