Eli Papa 17:21, 14 January 2008 (CST)
- 50 ug of antigen
- 200ul of final injection volume per mice
- 100ul of antigen + PBS
- 100ul of adjuvant
- allow some extra room for dead volumes in syringes, etc (prepare for 4 mice instead of 3, 8 mice instead of 6, etc)
NOTE: use complete Freud's adjuvant only in the primary injection. Any booster should be done by incomplete adjuvant or otherwise the 2ndary response may be lethal.
- load antigen + pbs in an eppendorf
- syringe adjuvant out of bottle (gauge 21 is recommended, nothing much thinner). DO NOT MIX at this point
- can put into eppendorf and then pipette to be more precise (allowing sufficient time for the pipette to aspirate the viscous material) or leave in the 3ml syringe.
- load into 3ml syringe using 21g
- connect syringe to 3-way (one screw), making sure the screw side is closed
- connect other syringe
- mix adjuvant + antigen by repeatedly pushing and pulling. (push with one, pull with the other)
- go to mouse room
- prepare your cage
- prepare injection
- push all the immunogen in one of the two 3ml syringes
- screw off the empty 3ml syringe
- loosen the 1ml syringe by moving piston up and down a couple of times
- plug the 1ml syringe, making sure you have a good hold and can apply pressure (there is no screw)
- while holding the 1ml syringe (otherwise it flies away), push immunogen into it from the 3ml syringe
- take the 1ml syringe off and cap with a 25g 5/8 (nothing thinner as the adjuvant does not flow much)
transfer all immunogen emulsion
- immobilize each mice and inject IP.
- generally, 10 days should pass after primary immunization. longer is fine, shorter (min ~ 7 days) can also work if one needs to rush. Consider that the peak cellular response occurs at days 4-7.
- booster injections can be done as close as 1 wk. from each other