Crosslinkers immunogenicity

Click chemistry or Staudinger ligation equipped heterobifunctional crosslinkers containing a hydrophilic PEG spacer provide a stable, yet highly efficient means to conjugate or immobilize biomolecules. The PEG spacer provides water solubility, low nonspecific binding character, and low immunogenicity. 

Figure 19.1 Immunogens are made by the crosslinking of a hapten molecule with a carrier using a conjugation reagent.

Figure 19.13 Conjugation to KLH often can cause precipitation due to the high-molecular weight of the carrier protein. The conjugation of [Met5]-enkephalin to KLH yields a soluble immunogen if the level of EDC addition is about 0.1 times that typically used with BSA as a carrier. This figure shows the gel filtration pattern on Sephadex G-25 after completion of the crosslinking reaction. The first peak is KLH and the second peak is excess peptide. Depletion of the peptide peak correlates to hapten-carrier conjugation.

Figure 19.14 Hapten-carrier immunogen conjugates can be formed using homobifunctional NHS ester crosslinkers. The reaction may create large polymeric complexes, some of which could precipitate.

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