I conjugated my 25 KD protein to Anti DEC-205 mAb by polysaccharide groups in Ab structure. I didn't have SMCC or Sulfo-SMCC then I used this protocol for conjugation. Now I have smears with high molecular weight ( > 200 KD). I must immunize my mice by this conjugated product. How much of this product is suitable for immunization? Which of them is important? The amount of protein in conjugated product or amount of whole conjugated product? In other words is it necessary to calculate the amount of protein in conjugated product? If i want to immunize my mice with 5 micrograms, this 5 micrograms is the net protein in conjugated product or is the amount of Ab-protein product? I sent the western-blot picture of my conjugated product.