Polyclonal antibodies arise from different B cells within the animal when the Immunogen injected. The polyclonal antibodies recognize different epitopes, (i.e. small regions within the antigen molecule). They could be considered as an assemblage of multiple monoclonal antibodies which against different epitopes. The advantage of using these antibodies in immunoassays lies in their ability to saturate the antigen, (i.e. more antibody molecules can bind to the antigen for enhanced detection). However, due to their wider range of specificities there can be more "nonspecific" binding. Depending on the assay design, this can result in higher background staining.
Monoclonal antibodies are the product of an individual B lymphocyte clone. Antibodies from a given clone are immunochemically identical and react with a specific epitope on the antigen against which they are raised. For reasons of economy, mice are currently used almost exclusively for the production of monoclonal antibodies. However, the secondary generation of rabbit monoclonal antibody developed in SinoBiological Inc. has many advantages when compared with mouse monoclonal antibody. We will introduce the technology in other webpages. In conventional hybridoma technology, after an immune response has been achieved, mouse B lymphocytes from spleen or lymph nodes are harvested and fused with non-secreting mouse myeloma cells under specific chemical conditions. While the B lymphocytes convey the specific antibody, myeloma cells bestow upon the hybrid cells (hybridoma) longevity in culture medium. Cells that are not hybridized soon die in culture. The antibody-producing hybrid cells are cultured and tested for desired reactivity. Cells producing antibody to the antigen are selected for propagation in culture medium or by transplantation into the peritoneal cavity of syngeneic mice. Antibodies can be harvested from the resulting ascites fluid. And also, the monoclonal hybridoma could use in vitro cell culture systems to produce monoclonal antibodies. Thus, large and, at least theoretically, unlimited quantities of monoclonal antibodies of specific characteristics can be produced.
In immunoassays, monoclonal antibodies have certain advantages over their polyclonal counter parts. These advantages include high homogeneity, high specificity, ease of characterization, and no batch-to-batch or lot-to-lot variability in affinity. However, some pitfalls in the use of monoclonal antibodies should be noted. All too often, monoclonal antibodies are characterized using fresh or frozen tissue. If the antibody is to be used for formalin-fixed specimens, the targeted epitope must survive fixation. In some cases, polyclonal antibodies that target multiple epitopes within a single antigen have a better chance of detecting that antigen than monoclonal antibodies that are restricted to a single epitope. However, some antigens may have two or more identical epitopes and are described as multivalent.
The targeted epitope must also be unique to a given antigen. Specificity, one of the greatest benefits of monoclonal antibodies, is lost if the antibody recognizes an epitope shared by two or more different antigens (Antibody Cross-Reactivity).
|Polyclonal Antibody||Monoclonal Antibody|
Research-based users are more prefer to utilize the monoclonal antibodies due to their higher specificity. However, there are many antibody manufacturers globally; the quality of the antibodies is diverse. Based on the feedback of users around the world, some monoclonal antibodies to hot target molecules had good performance characteristics, so that the clone was accepted and become a popular classic clone. Sino Biological Inc. (SBI) company has improved and reliable antibody production and quality control system, high-level professional and technical personnel, developed and produced various antibodies strictly followed the GMP standards for pharmaceutical companies. The vast majority of monoclonal antibodies were comparable to the classic clones, which had good reputation internationally, in homogeneity, specificity, sensitivity, affinity, etc..
For example, the conjugated MAb against human CD3E compared with CD3E MAb (clone 145-2C1, BD Biosciences):
|Antigen: Human CD3E
Antibody: CD3e / CD3 epsilon Antibody (FITC), Mouse MAb (10977-M001-F);
CD3e / CD3 epsilon Antibody (APC), Mouse MAb (10977-M001-A);
CD3e / CD3 epsilon Antibody (PE), Mouse MAb (10977-M001-P)
|Cell:Human whole blood lymphocytes
Flow Cyt:Use 1µl for 10^6 cells
Instrument: BD FACSCalibur
Flow cytometry is a method to evaluate cell membrane proteins and intracellular proteins as well as peptides and DNA. The principle behind FACS is an antigen-antibody reaction, with the antibodies being fluorescently labelled. There are three fluorescent proteins (R-PE, APC, and PerCP) conjugated to antibodies. Flow cytometry quantification is carried out with intercalating color labels (without the antibody). Flow cytometry antibodies are widely used in cell counting, cell sorting, biomarker detection and protein engineering.