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Home  >  Fusion Protein Antibody Protocols  >  Fusion Protein Protocol in Rats and Mice

84 Day Fusion Protein Protocol - in Rats and Mice

Our experience shows that for certain Fusion Protein Antigens, this protocol works well.  Please note that the route of immunizations is intraperitoneal and there is  one intravenous injection.  Please keep in mind for the IV injection, the antigen supply must easily pass through 23 gauge needle and be in PBS buffer.  This protocol is not suitable for antigen on gel.

Day 0 -

Preimmune Bleed, 0.1ml of sera. Preimmune bleed is stored until the first immune bleed is sent. 

Inject 50 to 100ug intraperitoneal in Complete Freund's Adjuvant (FCA).   Antigen supply should be 0.8mg/ml or higher.

Day 28 -

Inject 25 to 50ug intraperitoneal in Incomplete Freund's Adjuvant (IFA)

Day 42 -

Inject 10 to 25ug intraperitoneal with IFA.

Day 56 -

Bleed, about 0.1ml sera

Inject 12.5ug intraperitoneal with IFA.
Inject 12.5ug intravenous without adjuvant.

Day 63 -

Bleed.  This bleed will be sent to you along with the preimmune bleed and the day 56 bleed 

Day 70 -

Inject 10 to 25ug intraperitoneal with IFA

Day 84 - Select one of the three options below:

For CLOSED projects - Bleed at day 84 - rat or mouse is then euthanized / discard (lowest cost).  The bleed is then sent to you.

For CLOSED projects - The rat or mouse is exsanguinated (final bleed) at day 84. The exsanguination sera is then sent to you. 

For OPEN projects - Bleed - This bleed is then sent to you.

After Day 84 for OPEN projects -

You may select one of the project extensions below.  If you do not specify a project extension, we will automatically begin Extension A.  Keep in mind that all procedures and maintenance after day 84 will be charged according to the Custom Polyclonal Antibody Production Price Schedule.

Extension A:

We will boost every four weeks with 10-25ug of antigen intraperitoneal with IFA. We will do a large 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 rat or mouse requires us to stop production bleeds.  The volume of serum collected for large or production bleeds depends on the weight and the general health of the rat or mouse. Generally, bleed volumes will not vary from the previous bleed drawn.    During this time, we boost the rats and mice every four weeks with 10 to 25ug of antigen 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.  Please keep in mind, unlike our other species, rats and mice do not tolerate many cycles of production bleeds.

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. 

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, rat or mice should be boosted 10 to 21 days before exsanguination.
2.      Discard - termination without sera - the rat or mouse is humanely euthanized.
        
When requesting termination, you must confirm the species and animal number. Written confirmation by email may be requested for terminal procedures.

Custom Extension:

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