Polyclonal Antibodies and Monoclonal Antibodies

Polyclonal antibodies (pAb) can recognize multiple epitopes on any antigen, while monoclonal antibodies (mAb) only detect one epitope on any antigen (as shown in the figure below). However, both polyclonal antibodies and monoclonal antibodies have their own advantages and disadvantages. The differences between polyclonal antibodies and monoclonal antibodies are discussed below:

Advantages of polyclonal antibodies:

1. Polyclonal antibodies can help to amplify the signal of a target protein at low expression levels, because the target protein can bind more than one antibody molecule on multiple epitopes. But this will adversely affect quantitative experiments as the results will become inaccurate.

2. Polyclonal antibodies give better results in IP/ChIP due to recognition of multiple epitopes.

3. Smaller changes in antigen are tolerated better than monoclonal antibodies.

4. They identify proteins with high homology to immunogenic proteins or screen for target proteins from tissue samples of non-immunogenic species, for example polyclonal antibodies are sometimes used when the nature of the antigen in the untested species is unknown. This also makes it very important to test the immunogenic sequence to determine if there is any cross-reactivity.

5. Polyclonal antibodies are usually the first choice for detection of denatured proteins.

6. Multiple epitopes generally provide more robust detection.

7. Polyclonal antibodies are not suitable for probing specific domains of an antigen because antisera often recognize multiple domains.

 

Disadvantages:

1. Prone to batch-to-batch variation.

2. Generates large amounts of non-specific antibodies, which can sometimes cause background signal in some applications.

3. Multiple epitopes make it important to examine immunogenic sequences to determine if there is any cross-reactivity. 

 

Objective facts:

1. Recognize multiple epitopes on any antigen. The resulting serum will contain a heterogeneous complex mixture of antibodies of different affinity.

2. Polyclonal antibodies are mainly composed of the IgG subclass.

3. Peptide immunogens are commonly used to generate polyclonal antibodies targeting unique epitopes and are particularly suitable for families of proteins of high homology.

 

Antibody preparation:

1. The technology and skills required for preparation are not high.

2. Preparation time is short.

3. Polyclonal antibodies are not suitable for probing specific domains of an antigen because polyclonal antisera often recognize multiple domains.

 

Advantages of monoclonal antibodies:

1. Once a hybridoma is made, it becomes a constant source of regeneration and will be the same for all batches – very helpful in ensuring consistency and standardization of experimental procedures and results.

2. Monoclonal antibodies generally cause low background in sections and cell staining. Because they detect one target epitope with greater specificity, they are less likely to cross-react with other proteins.

3. Due to their specificity, monoclonal antibodies are well suited for use as primary antibodies in analytical assays, or for the detection of antigens in tissues, and typically produce significantly lower background staining than polyclonal antibodies.

4. Monoclonal antibodies are very homogenous compared to polyclonal antibodies. Monoclonal antibody results are highly reproducible across experiments if the experimental conditions are kept constant.

5. The specificity of monoclonal antibodies enables them to bind antigens efficiently in mixtures.

 

Disadvantages:

1. A large number of specific antibodies can be produced, but may be too specific.

2. More prone to loss of epitopes after chemical treatment of antigens than polyclonal antibodies. This can be compensated by using two or more monoclonal antibodies to the same antigen.

 

Objective facts:

1. Only one epitope on the antigen is detected.

2. They consist of only one antibody subtype. When a secondary antibody is required for detection, the antibody should be selected against the correct subclass.

 

Antibody preparation:

1. High technology is required.

2. Training is required before using the technology.

3. The preparation of hybridomas takes a long time.