A Short Guide to Antibody Production

Posted by eliteinternet on October 25th, 2019

Before beginning, some definitions will help the laymen to understand the variety of products available on the market today: 

Polyclonal antibodies are usually affinity purified.  As such, the antibody mixture (not the host animal) is exposed to the antigen.  A polyclonal antibody consists of agroup of immunoglobulins, generated from a B-cell clone. The antibodies search epitopes, also known as binding sites, on a single antigen. The derivation from various B-cell clones and subsequent targeting of several epitopes sets polyclonal antibodies apart from their monoclonal counterparts. 

The LIMK1gene encodes the enzyme LIM domain kinase 1 in humans. 

GCN2 is a serine/threonine-protein kinasethat senses amino acid deficiency through binding to uncharged transfer RNA (tRNA). The protein assists in modulating amino acid metabolism in response to nutrient deprivation. 

How Polyclonal Antibodies are Made 

Polyclonal antibodies are made in a process that elicits an immune response called immunization.  The host receives an injection, and its immune system recognizes the injection as a foreign substance, prompting antibodies that cause immune blockage or clearance.  If an immune system response is not mounted, the use of adjuvants is necessary.  Such adjuvants include Freund's adjuvant or keyhole limpet hemocyanin (KLH).  This process further drives immune recruitment. Immunization largely consists of a process used to create polyclonal antibodies. 

The first host exposure to antigen is called the "Prime" immunization. It begins with the exposureof an injection into the skin, peritoneum, subcutaneous layer, or muscle. Oral and intranasal administration are less commonly used.  Additional exposure to the antigen is called a "boost" immunization. 

In order to evaluate the progress, it is important to measure the antibody levels during the immunization process.  ELISA against target antigen is used to compare immunized serum samples against pre-immunized samples to show the degree of immune response as a result of the immunization. In general, a 1:64,000 titer is believed a strong immune response and appropriate for proceeding to lymphocyte fusion or serum harvest. If the titer is too lower, subsequent boost immunizations, alternative routes of administration, or additional adjuvant may be required. 

Serum is utilized as a molecular biology probe.  However, it often undergoesadditional purification to increase sensitivity and specificity of an assay. Affinity purification leads to antibody recovery to different degrees based on the chosen technique.