Category Archives: antibody function

in this part , we mainly introduce the function of antibody

The Difference Between Antigen and Antibody

Summary

Sometimes you may know what is antibody and what is antigen. However, can you be sure that you are able to point the difference between antigen and antibody? If not, you can find the answer in this article.

antigen and antibodyBody

What is antigen

Antigen is a substence, which can stimulate the immune system and induce an immune response. Such as antibodies or sensitized lymphocytes specifical reaction. A complete antigen should be including two parts: immunogenicity and immunoreactivity. The first refers to the substance, which is capable of inducing a host cell to produce an immune response; the second one is also known as reactogenicity, it refers to the antigen, which can react with antibody or sensitized lymphocytes. Some substances, which alone can just have the reactogenicity no immunoreactivity, are called hapten; however immunogen usually has both immune response capability.

What is antibody

Antibody is always called immunoglobulin(Ig). It refers to the immunoglobulin, which have antibody activity and antibody-like structure. Ig, which is about 20% of plasma protein, is produced by plasma cells, existing in blood and other body fluids. Ig structure has heterogeneity, it can be divided into different types; A majority of Ig have antibody activity, they can specifically recognize and bind antigen, and produce a series of biological reactions.

Immune functions of the antibody

Antibody binds to a specific antigen to occur biological effects, the ultimate physiological function is mainly anti-infection, the mechanism can be classified as following aspects:

  1. Toxin neutralization

When the antigen is a bacterial toxin, it binds to the antibody, then they can neutralize toxicity of the host cell. When the antigen is a hormone or enzyme, it binds to the antibody, so it may also be losing its toxicity.

  1. Infection of neutralization

When the virus bind to the corresponding antibody, it may lose its ability to invade into host cells to reproduce.

  1. Phagocytosis conditioning

After antibacterial antibody combined with bacteria, although it can not directly eliminate the cells, being used as the conditioning factor, which promotes phagocytosis of immune cells to engulf bacteria.

  1. Induced lysis

After IgG antibody and IgM antibody bind] to bacteria, they can activate the classical complement activation to lysis pathogens.

  1. Antibody-mediated ADCCIgG class can also mediate NK cells produced to ADCC effector, then it can kill virus-infected target cells and malignant cells.

All in all, sometimes antigen and antibody are very hardly to be separated. We can just tell them from each other by their origins.

These are the main defferences between antigen and antibody. I am not sure if this is helpful for you. But I can be sure that this must let you know more about the two substances.

 

Antibody Engineering and Antibody Engineering Drugs

antibody engineering drugsSummary

In the 21st century, biotechnology and information technology is providing a strong impetus for global economic development. Antibody engineering technology with the development of modern biotechnology and continuous improvement, is the main force of the biotechnology industry, in particular, plays an important role in the field of bio-pharmaceutical technology. In this article, I will introduce something about antibody engineering and its drugs.

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What is antibody engineering

Antibody engineering is the engineering to create the antibody molecules. It is depending on using recombinant DNA and protein engineering technology to modify and antibody gene, after transfecting the proper recipient cells, it use the antibody molecule expression, or cell fusion and chemical modification methods to achieve its goal.

All kinds of antibody engineering

Antibody engineering includes antigens, antibodies, polyclonal antibodies, cell engineering, genetically engineered antibody, purification and identification of antibodies, antibody technology applications and immunological tests

What is antibody engineering drugs

antibody engineering drugs, also called antibody drugs, Monoclonal antibody therapy agent or antibody therapy agent, is used to cure monoclonal antibody, Antibody fragments, antibody improved by gene engineering and Antibody conjugates or Antibody fusion protein.

The importance of antibody engineering drugs

Malignant, the biggest enemy for human health. It is the core cause of human death. The traditional therpy of the it are Surgery, radiotherapy and chemotherapy. Recent years, the development of monoclonal antibody drugs has stepped into another stage. Monoclonal antibody can recognize malignant and can effectively improve the lethality of drugs, reduce side effects. Therefore, it attract more attention from people.

In the early 1980s, the research antibody gene structure and function of recombinant DNA technology combined to produce a genetically engineered antibody technology. Then genetically engineered antibody will process the antibody gene according to the different needs, and then introduced into an appropriate recipient cell to express. Genetically antibody engineering includes chimeric antibody, humanization(human reshaped antibody or CDR shift, epitope guided selection EGS and etc.), small monoclular antibody, multivalent and bispecific antibodies and antibody fusion protein

Compared with monoclonal antibody, genetically engineering antibodies have the advantages as follows:

  • Through the transformation of genetic engineering techniques, we can reduce or even eliminate rejection of human antibodies.
  • Genetically engineered antibody of smaller molecular weight can be partially reduced murine antibody, and it is more conducive to penetrate the vessel wall into the core of the lesion site
  • According to the need of treatment, we can prepara new antibodies

This is the brief introduction to antibody engineering and antibody engineering drugs. There are much other details, which is not mentioned. You can find it by yourself.

Something You do not Know About Chimeric Antibody

Something You do not Know About Chimeric Antibody

Summary

In the antibody field, what familiar to us are the Genetically engineered antibodies and Monoclonal antibody. However, one of the most important antibody we do not know is the chimeric antibody. In this article I will introduce some knowledge about this antibody.chimeric antibody

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Definition

Chimeric antibody was the first success in the preparation of genetically engineered antibody. It is composed of V region gene and gene-splicing Cregion of a human antibody. And then it is inserted into a vector, transfection of antibody molecules myeloma tissue expression.because it reduces the murine components, thereby reducing the adverse effects caused by murine antibody, and helps improve the outcome.

The features of chimeric antibody

  • It not only retains the parent murine antibodies with high specificity and affinity, but also reduces the murine component about 70%.
  • The human Fc portion can effectively mediate biological effector functions, such as antibody-dependent cellular cytotoxicity mediated (Antibody dependent cell-medi-ated cytotoxicity, ADCC), complement-dependent cytotoxicity (Complement dependent cytotoxicity, CDC ), etc.
  • It is free to choose the type of antibody, subtypes, classes, subclasses, size, domain and add glycosylation sites, etc., in order to take advantage of their different physical and chemical properties and biological characteristics;
  • Chimeric antibodies Expression “box”, which is produced by efficient eukaryotic expression vector and a human antibody, allows the insertion of different variable region of the murine monoclonal antibody, shortens the operation and development cycle;

The steps to produce chimeric antibody

Production of chimeric antibodies, the first step is the key step is to obtain from hybridoma cell McAb specific variable region gene, and then connect with the human antibody constant region genes, and finally to build a good human – mouse chimeric antibody gene expression in myeloma cells. The main steps include murine antibody variable region gene cloning, fishing human antibody constant region gene extraction, vectors and host cells are constructed gene splicing human antibody constant region gene of murine antibody variable region and as well as efficient expression screening positive cells and expression identification products

  • Cloning of murine antibody variable region gene
  • Human antibody constant region gene angling
  • Human – mouse chimeric antibody expression vector
  • Efficient expression vectors and host cells Construction
  • Human – mouse chimeric antibody expression and identification

Chimeric antibody, as the first born of genetically engineered antibody, has its unique advantages: retaining parental murine antibodies with high specificity and affinity; having a longer half-life in the body; playing a variety of biological functions effectively; with easy operation and low cost and great prospects for clinical application etc.

A Brief Look on Protein Sequencing

A Brief Look on Protein Sequencing

Tags: protein sequencing

protein sequencingSummary

We talk about the protein sequencing is mainly about the primari structure of protein sequencing. This primary structure includes the the number of polypeptide chains in protein. And in many cases, polypeptides and protein can be used as the one. The oder of the acid amino in polypeptide is the bio-functional bases of protein.

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Requirment of the sequencing

Sample purity(over 97%)

Accurate molecular weight of proteins

Accurate number of subunits

Determination of amino acid composition of the protein; and calculate the number of each amino acid in the molecular weight.

Determination of the amount of ammonia in the hydrolysis, calculated amide content

Determination of protein and amino acid sequence of a polypeptide

Open and isolation of peptide chain

The number determination of protein molecules in the polypeptide chain

Break of disulfide bonds

Determination of the amino acid composition of each polypeptide chain, and calculate the molecular ratio of amino acid components

Determination of the N-terminal, C terminal

Polypeptide chain scission

Determination of the amino acid sequence of each peptide.

The steps of sequencing

  1. Split the polypeptide chain

Protein molecules are composed of multiple polypeptide chains, it must be firstly splited. Several polypeptide chains are linked together with the help of non-covalently, which is known as oligomeric proteins.

  1. To determine the number of the polypeptides in protein.

Determination of the number of the polypeptide chain in protein molecules. Through the determination of amino acid residues at the end of the number of moles and the relationship between the protein molecular weight, can determine the number of the polypeptide chain..

  1. Disulfide bond rupture.

Several polypeptide chain through disulfide bond together, can be in 8 mol/L urea or, in the presence of 6 mol/L guanidine hydrochloride with excessive  – mercaptoethanol processing, make the disulfide bond reduction of sulphur, and then use generated thiol alkylating reagent protection, to prevent the oxidation of the it to be.

  1. To determine the amino acid composition, each of the polypeptide chain and calculate the amino acid composition of molecular ratio
  2. Analyzing the N – end of a polypeptide chain and C – end
  3. Polypeptide chain rupture into multiple peptides. Two or more different fracture method can be used to peptide broken into two or more sets of peptides or peptide fragments, and the separation
  4. The determination of each amino acid sequence of peptides
  5. Determine the peptides sequence in the polypeptide chain. To use two or more sets of amino acid sequence of peptides crisscross overlap each other, piecing together the whole amino acid sequence of a polypeptide chain.
  6. To determine the location of disulfide bond in the polypeptide chain

Those are the steps of protein sequencing. And you can take those tips to achieve your goals.

You Know How antibody generate?

You Know How antibody generate?

Topic covered

Summary

Discovery

Physical and chemical properties

Biological Activity

Production

Classification

Summary

Antibody, an immunoglobulin, is generated by the B cells in the antigenic substance’s stimulation. And it can be reacting with special antigen. Antibody molecule are synthesized and secreted by the plasma cells, and each can produce a plasma cell clone specific antibody molecules, so the antibody in serum is mixed by many kinds of antibody molecules.

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Physical and chemical properties of antibody

In the first 40, Tiselius and Kabat had proved the activity of antibody has something to do with the Globulin in serum. They immunized rabbit with the pneumatically and polysaccharide to get the high-quality immunological serum. So they proved that the activity of antibody has something to do with the Globulin.

The biological activity of the antibody

  1. Specific antibodies and antigen binding antibody production stimulating substance are called an antigen, an antibody and its corresponding antigen molecule binding is called specific binding.
  2. Binding of antibodies and complement, under certain conditions, the antibody molecule may be presented in the serum complement molecules combination and make it activate, produce a variety of biological effects, complement binding phenomenon and it is known as antibody, antibody molecules revealed inter-molecular interactions complement.
  3. The third function of antibodies is antibodies can enhance phagocyte role. In experiments in vitriol, the serum added suspension of necrophiliacs and enhancing phagocytes corresponding cells is called as an antibody conditioning.

The production of antibody

In order to study on the Physical and chemical properties, molecular structure and function, and the application of antibodies in clinical diagnosis, treatment and prevention of antibody, we should produce antibody by human beings. Recently, according to the methods and the rules of preparation, we classify the antibody into three kinds: poly-clonal antibodies, monoclonal antibodies and genetically engineered antibodies.

  1. Poly-clonal antibody

The early conventional antibody preparation method is a natural antigen immunized animals by various routes, as antigenic material with multiple epitopes, it can stimulate the production of a variety of antibody-forming cell clones, synthesis and secretion against various epitopes antibody secreted into the serum or body fluids, it is in fact a mixture of the serum with a variety of antibodies, such known serum by in vivo immunization of the obtained poly-clonal antibody, it is the first generation of antibodies.

  1. Monoclonal Antibodies

Immune method is difficult to obtain a monoclonal antibody (monoclonal antibody, McAb). The antibody can require forming cells and then it can be selected and cultured to obtain a monoclonal antibody specific of known in vitriol. 1975 German scholar Kohler and Milstein from British got the mouse melanoma cells by sheep red blood cells to immunize mouse splenocytes in vitriol fusion of two cells, and found that both the partially formed hybrid cells can continue in vitriol culture conditions growth and reproduction can secrete anti-SRBC antibody, this is called hybrid hybridism cell line.

3.     Genetically engineered antibodies

Since 1975, the advent of monoclonal antibody hybridism technology, monoclonal bodies in medicine is widely used in the diagnosis and treatment of diarrhea. But the vast majority of the murine monoclonal antibody, when administered in vivo to produce clinical repeated anti-mouse antibody, causing the clinical efficacy diminished or disappeared. Therefore, the clinical application of the ideal of a monoclonal antibody should be of human origin, but the human – human hybridism technology has yet to breakthrough, even if successfully developed, there are also people – human hybrid tumor passages unstable, low antibody affinity and production is not Higher problem. Office failed to present a better solution is to develop genetically engineered antibody, (genetically engineering antibody) in place of the murine monoclonal antibody for clinical use.

This is the brief introduction of the antibody’s origin and production.

Polyclonal Antibodies and Monoclonal Antibodies—-Which is More Powerful?

Polyclonal Antibodies and Monoclonal Antibodies—-Which is More Powerful?

Topic covered:

Introduction

Form of comparision

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Introduction

Monoclonal antibodies are produced by a specific antibody. Monoclonal antibodies can be targeted with a sigle specific epitope binding, it is just like the missile hit the target exactly. On the other hand, even if the same eoitope, the clones can generate several kinds of antibodies in bodies, then it forms a mixture of monoclonal antibodies, which is called polyclone antibodies.

Polyclonal antibodies Monoclonal antibodies
Low cost High cost
General techniques High techniques
Short time cost Long time needed for hybridomas
A large number of non-specific antibody A large number of non-specific antibody with strong specificity
Many epitopes on any antigens One epitope on one antigen

Form

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Polyclonal antibodies

Advantages show

 

  1. Polyclonal antibodies may help amplify low level of expression of the target protein signals, since the target protein can bind more than one antibody molecule in multiple epitopes. But this will would adversely affect the quantitative experiments, because the result will be inaccurate.
  2. Because the polyclonal antibodies can recognize multiple epitopes, it can get better results in IP / ChIP.
  3. Polyclonal antibodies can give much more space for the antigens than the monoclonal antibodies.
  4. Polyclonal antibodies can recognize the protein, which has high homology to immunogenic protein. Or it can tissue sample from a non-immunogenic target protein screened species. This is also significant to make the detection immunogen sequence to determine whether there is any crossreactivity.
  5. Polyclonal antibodies are generally preferred to detect denatured proteins.
  6. Multi-epitope generally offer more powerful detection.

 

Disadvantages show

  1. Polyclonal antibodies prone to differences between batches.
  2. Because antisera usually recognize multiple domains, polyclonal antibodies are not suitable for detecting antigens for specific domain.
  3. It will cause a large amount of non-specific antibodies, sometimes the background signal in certain applications.
  4. Multi-epitope is very important for detection of the immunogenic epitope sequence, which can be used for determining whether there is any cross-reactivity.

 

Objective fact

  1. To identify any multiple epitopes on an antigen. And the sera obtained will contain the mixture of complex heterogeneous, which is generated by the differen affinity antibodies.
  2. Polyclonal antibodies are mainly formed by IgG subclasses.
  3. Peptide immunogens is commonly used for targeting a unique epitope polyclonal antibodies, especially for highly homologous protein family.

 

Antibody preparation

  1. Low production cost
  2. Preparation techniques and skills needed are not high.
  3. Short preparation time

 

Monoclonal antibodies

Advantages show

  1. After prepared, hybridomas became a constant source of regeneration. All batches will be the same—to ensure constency and standardization eexperimental procedure and to be very helpful to the resulte.
  2. Monoclonal antibodies are usually cause the low background in the slice and cell staining. Because they are more specific detection of a target epitope. So it is less likely to cross-react with the other proteins.
  3. Because of specificity, monoclonal antibodies are very suitable for using in an anti-determination, or for detection of the antigen in cells, and the background staining is lower than polyclonal antibodies.
  4. Compared with polyclonal antibodies, monoclonal antibodies have high homogeneity. Within the same condition of experiment, the results of experiment with monoclonal antibodies has high reproducibit]lity.
  5. The specificity of monoclonal antibodies can make it bind with antigen in the mixture.

Disadvantages show

  1. It can generate a large amount of specific antibodies, but the specificity may be too strong.
  2. It is much more easier than polyclonal antibodies in lossing epitope, after chemical treated by antigens.

Objective facts

  1. It only detects one epitope on antigens.
  2. It is only formed by one antibody subtypes. When we need secondary antibody to detect antibodies, we should select the correct subtypes.

Antibody preparation

  1. High techniques are required.
  2. It can be used after preparation.
  3. Long time cost for hybridomas

These are comparation between polyclonal antibodies and monoclonal antibodies. We can not tell which one is powerful, because each one of them has its own advantages.

Monoclonal antibody

Monoclonal antibody

Monoclonal antibodies (mAb or moAb) are monospecific antibodies that are made by identical immune cells that are all clones of a unique parent cell, in contrast to polyclonal antibodies which are made from several different immune cells.

Monoclonal antibodies have monovalent affinity, in that they bind to the same epitope.

Given almost any substance, it is possible to generate monoclonal antibodies that specifically bind to that substance; they can serve detect or purify that substance. So this has been an significant tool in biochemistry, molecular biology and medicine. The non-proprietary drug name ends in–mab, when it is used as medications, and many immunotherapy specialists use the word mab anacronymically.

Now, we will introduce the production of the monoclonal antibody.

if we need an antibody for avian SERCA2 protein, we would inject the protein into a mouse. This is typically done in two doses, an initial “priming” dose and a second “booster” dose 10days later.
monoclonal antibody

The production of monoclonal antibodies was pioneered by Georges Kohler and Cesar Milstein in 1975. Let us see how their method, now tried and tested for over 20 years, would be applied in a particular case. In order for us to isolate a B lymphocyte producing a certain antibody,we first have to include the production of such a B cell in an organism. For instance, if we need an antibody for avian SERCA2 protein, we would inject the protein into a mouse. This is typically done in two doses, an initial “priming” dose and a second “booster” dose 10days later. Since the protein is of foreign origin, the mouse immune system recognizes it as such and soon some of the B cell in the mouse would begin production of the antibody to avian SERCA2.

The uses of monoclonal antibodies

Monoclonal antibodies have a variety of academic, medical and commercial uses. It would be impossible to list all of these here. But the following list should indicate how ubiquitous monoclonal antibody technology has become in biotechnology.

  • Antibodies are used in several diagnostic tests to detect small amounts of drugs, toxins or hormones, eg monoclonal antibodies to human chorionic gonadotropin (HCG)are used in pregnancy test kits. Another diagnostic uses of antibodies is the diagnosis of AIDS by the ELISA test. (Biotech, 1989)
  • Antibodies are used in the radioimmunodetection and radioimmunotherapy of cancer, and some new methods can even target only the cell membranes of cancerous cells. (Chaudhari et al, 1994) A new cancer drug based on monoclonal antibody technology is Ritoxin, approved by the FDA in November 1997(Orrs, 1997)
  • Monoclonal antibodies can be used to treat viral diseases, traditionally considered “untreatable”. In fact, there is some evidence to suggest that antibodies may lead to a cure for AIDS (P/S/L, 1997)
  • Monoclonal antibodies can be used to classify strains of a single pathogen, eg Neisseria gonorrhoeae cna be types using monoclonal antibodies (Wang et al, 1977).

 

This is the function and uses of the monoclonal antibodies.

if we need an antibody for avian SERCA2 protein, we would inject the protein into a mouse. This is typically done in two doses, an initial “priming” dose and a second “booster” dose 10days later.
monoclonal antibody