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Home  >  PRF&L Traditional Protocols  >  Rat and Mouse custom antibody production

TRADITIONAL 70 DAY POLYCLONAL ANTIBODY PRODUCTION - in Rats & Mice


Day 0 -

Preimmune Bleed. Preimmune bleed is stored until the first immune bleed is sent.  Rat preimmune bleed generally yields 0.2ml plasma, mice 0.1ml.

Inject 50ug Intraperitoneal Complete Freund's Adjuvant (CFA).   We will inject intraperitoneal and subcutaneous if the concentration of antigen is less than 0.5mg/ml. Acrylamide gel can only be injected subcutaneous. .

Day 14 -

Inject 25ug intraperitoneal in Incomplete Freund's Adjuvant (IFA)

Day 28 -

Inject 25ug intraperitoneal with IFA.

Day 42 -

Bleed.  0.2ml plasma for rats, 0.1ml plasma for mice. We will ship via Federal Express the preimmune bleed and this first immune bleed.  It is then up to the investigator to test the plasma, determine the quality of antibodies, and let us know if any changes are necessary in the protocol.

Day 56 -

Inject 15ug intraperitoneal with IFA

Day 70 -

Bleed.  0.2ml plasma for rats, 0.1ml plasma for mice. We will ship via Federal Express this small bleed. It is up to the investigator to test the plasma, determine the quality of antibodies, and let us know if any changes are necessary in the protocol.

After Day 70 -

You may select one of the project extensions below.  If you do not specify a project extension, we will automatically begin Extension A.

Extension A:

We will boost every four weeks with 10 to 15ug of antigen subcutaneous or intraperitoneal with IFA. We will do a bleed two weeks after each boost and ship. We will continue to boost and bleed every four weeks until you contact us. 

Extension B:

When the antisera contains sufficient antibody titer and large volumes of sera are needed, we can start production bleeds. We do production bleeds every seven days.   We will continue with production bleeds until you contact us or the health of the rats or mice requires us to stop production bleeds.  The volume of plasma collected for production bleeds depends on the weight and the general health of the rat or mouse. General yield is 0.2ml to 0.25ml plasma per rat and 0.1ml plasma per mouse.    During this time, we boost the rats or mice with 10 to 15ug of antigen subcutaneous or intraperitoneal with IFA.  You may also request that we not do any boosts after the initial immunization scheme outlined above since some antigens and some animals may respond negatively to boosts during the period of peak antibody activity. Boosts may be required only when titer drops. It is up to the investigator to decide and let us know if we should discontinue boosts as our standard schedule provides for boosts every 28 days.    Shipping for production bleeds is every four weeks unless you request otherwise.  Unlike larger animals, production bleed cycles may not be able to be repeated more than twice in rats and mice.

Extension C:

Some investigators prefer to "wait and see." after day 70.  Extension C is hold without boosting and bleeding.  We will hold until you tell us to do something else.  Our experience shows that the peak antibody titer occurs at an average of 120 days, however we have experienced the onset of peak activity as early as six weeks and as long as a year. If results are not as you expect, please consult with us and we can make recommendations on how to change your protocol.

Termination:

When you have collected enough antisera, you will need to contact us to terminate the project. You may request that we terminate by:
1.      Exsanguination - termination with sera - the rat or mouse is anesthetized and the terminal bleed is taken.  For best results, rats and mice should be boosted 10 to 21 days before exsanguination. 
2.      Discard - termination without sera - the rat or mouse is humanely euthanized.
        
Many investigators request that we harvest the spleen from mice and rats at the time of termination.  Please contact us well in advance with a request for spleen harvest.  Several techniques are available - please let us know your requirements.

When requesting termination, you must confirm the species and animal number. Written confirmation by email or fax may be requested for terminal procedures.

Custom Extension:

You may design your own project extension.  Let us know what we can do for you!