# Design of Antibody-Functionalized Polymeric Membranes for the Immunoisolation of Pancreatic Islets
## 4 Conclusions  
In order to reduce the immunosuppression response in patients affected by diabetes 1 and transplanted with the pancreatic islets of a donor, several coating procedures of the donor islets have been proposed. Here we designed an immuno-functionalization strategy aimed to conjugate a therapeutically relevant antibody to PEG encapsulated islets. We proposed an approach based on the site-directed functionalization of the antibody while keeping functional the antigen-recognition site and thus preserving its functionality. Gold-coated silicon wafers were functionalized using 11-Mercaptoundecanoic acid and were used as a substrate for further stepwise modification, leading to a nickel(II)-terminated ligand surface. As a proof of principle, the specific immobilization was demonstrated for a generic fluorescent antigen antibody pair (GFPA-Anti-GFP), confirming the attachment of the His-tag protein to the solid support. The immobilized His-tagged protein was recognized by the target antibody, thus blocking the binding site; then the constant fraction of the antibody was functionalized with Acryl-PEG-NHS. The recovery efficiency of the PEG-antibody complex is not satisfying (less than 10% of the initial amount of antibody), thus requiring further implementation and modification of the protocol steps. Nevertheless, this approach proposes a controlled functionalization chemistry to preserve the binding site of the antibody. Moreover, murine pancreatic islets were successfully encapsulated with the PEG hydrogel bearing the Acryl-PEG-Anti-GFP complex, and their viability and functionality was not affected by the PEG coating. To show the general applicability of the approach, the immobilization protocol was extended was to a therapeutically relevant antigen-antibody couple, B7-1-CTLA4-Ig [25]. Even in this case, the recovery efficiency was quite low, suggesting that this methodology requires further optimization prior to become applicable in vivo. In conclusion, this strategy represents an attractive ready-to-use tool for customizing a patient-specific immunotherapy, by adjusting the specificity of the conjugated immunotherapeutic agent, based on an autoimmune response. The application of this methodology to the immunocoating of pancreatic islets deserves research efforts to optimize the quantitative yield of the process. Figure 12: Western blot analysis (**a**) and relative density percentage (**b**) of mother solution of CTLA-4 Ig (dark blue bar), CTLA-4 Ig recovered after incubation (light blue bar) and Acryl-PEG-CTLA-4 Ig complex eluted from the solid substrate (red bar). Reported values have been averaged over three measurements. Error bars represent standard deviation (SD) (*** \(p<0.001\), *** \(p<0.01\)). **Supplementary Materials:** The following are available online at [[http://www.mdpi.com/2076-3417/10/17/6056/s1](http://www.mdpi.com/2076-3417/10/17/6056/s1)]([http://www.mdpi.com/2076-3417/10/17/6056/s1](http://www.mdpi.com/2076-3417/10/17/6056/s1)), Figure S1: Values of contact angle of either gold surface, gold surface after 12 h and 60 h incubation with MUA, or gold surface after 60 h incubation with MUA and NTA. Reported values have been averaged over 5 measurements. Error bars represent standard deviation (SD). (*** \(p<\) 0.001, ** \(p<\) 0.01). Figure S2: Schematic representation of the experimental design for the conformal encapsulation of islets through photopolymerization. Islets are incubated with the photoinitiator that adsorbs on the islet surface. The islet-photoinitiator complex is incubated with the PEGDA and exposed to UV to encapsulate the islet, inducing the crosslinking of the polymer. **Author Contributions:** Conceptualization, L.B., A.Q., and M.M. ; methodology, A.C., U.M., A.B., and T.V. ; writing and editing, A.C., U.M., L.B., and A.Q. ; supervision, A.S. All authors have read and agreed to the published version of the manuscript. **Funding:** This research was partly funded by the Progetto regionale Lab on a Swab (cod. OTHZY54; funds to L.B.