在一个月内制备高效多克隆抗体的方法
丁香园论坛
1273
本人利用该方法制备了高效价的多克隆抗体，与大家分享！
Animals
Healthy male New Zealand rabbits aged 3-6 months (body weight ~ 2.5 kg) lived in animal house at least for 3-4 days for adapting to the new circumstance.
Polyclonal antisera production
Sample A: Emulsify your aimed protein 200-300 mg in PBS with same volume of the Freund's complete adjuvant (3-4 mg/ml heat-killed Bacillus tubercle in lanolin : mineral oil=2:5 [v/v] ) in 1 ml total volume.
Sample B: Same as injection solution I but no Bacillus tubercle.
The modified immunization method: At the first day, 1.5ml blood was collected by bleeding rabbits from the marginal vein of the ear for preparing the pre-immunized-sera as negative control. Afterwards, 1 ml of sample A in 30-50 ml per each site was injected intradermally on the back and proximal limbs of the rabbit[1, 3, 4]. On the 3rd day, repeat the above injection once to strengthen the first immunization stimulation. On the 28th day, 1 ml of sample B was injected by the same way. On the 35th day, 0.5 ml blood was collected by the same way used before for checking its titer of the antisera by ELISA. Harvest the antisera from the arteriae carotis.
Traditional immunization method: Exactly same as the modified method, except no additional sample A injection on the 3rd day. Besides, it needed several sample B injections in every 2-4 weeks successively.
Animals
Healthy male New Zealand rabbits aged 3-6 months (body weight ~ 2.5 kg) lived in animal house at least for 3-4 days for adapting to the new circumstance.
Polyclonal antisera production
Sample A: Emulsify your aimed protein 200-300 mg in PBS with same volume of the Freund's complete adjuvant (3-4 mg/ml heat-killed Bacillus tubercle in lanolin : mineral oil=2:5 [v/v] ) in 1 ml total volume.
Sample B: Same as injection solution I but no Bacillus tubercle.
The modified immunization method: At the first day, 1.5ml blood was collected by bleeding rabbits from the marginal vein of the ear for preparing the pre-immunized-sera as negative control. Afterwards, 1 ml of sample A in 30-50 ml per each site was injected intradermally on the back and proximal limbs of the rabbit[1, 3, 4]. On the 3rd day, repeat the above injection once to strengthen the first immunization stimulation. On the 28th day, 1 ml of sample B was injected by the same way. On the 35th day, 0.5 ml blood was collected by the same way used before for checking its titer of the antisera by ELISA. Harvest the antisera from the arteriae carotis.
Traditional immunization method: Exactly same as the modified method, except no additional sample A injection on the 3rd day. Besides, it needed several sample B injections in every 2-4 weeks successively.
