**Tomonori Waku \*, Saki Nishigaki, Yuichi Kitagawa, Sayaka Koeda, Kazufumi Kawabata, Shigeru Kunugi, Akio Kobori and Naoki Tanaka**

Faculty of Molecular Chemistry and Engineering, Kyoto Institute of Technology, Gosyokaido-cho, Matsugasaki, Sakyo-ku, Kyoto 606-8585, Japan

**\*** Correspondence: waku1214@kit.ac.jp; Tel.: +81-75-724-7811

Received: 22 May 2019; Accepted: 25 July 2019; Published: 2 August 2019

**Abstract:** Recently, nanofibers (NFs) formed from antigenic peptides conjugated to β-sheet-forming peptides have attracted much attention as a new generation of vaccines. However, studies describing how the hydrophilic-hydrophobic balance of NF components affects cellular interactions of NFs are limited. In this report, three different NFs were prepared by self-assembly of β-sheet-forming peptides conjugated with model antigenic peptides (SIINFEKL) from ovalbumin and hydrophilic oligo-ethylene glycol (EG) of differing chain lengths (6-, 12- and 24-mer) to investigate the effect of EG length of antigen-loaded NFs on their cellular uptake, cytotoxicity, and dendritic cell (DC)-stimulation ability. We used an immortal DC line, termed JAWS II, derived from bone marrow-derived DCs of a C57BL/6 p53-knockout mouse. The uptake of NFs, consisting of the EG 12-mer by DCs, was the most effective and activated DC without exhibiting significant cytotoxicity. Increasing the EG chain length significantly reduced cellular entry and DC activation by NFs. Conversely, shortening the EG chain enhanced DC activation but increased toxicity and impaired water-dispersibility, resulting in low cellular uptake. These results show that the interaction of antigen-loaded NFs with cells can be tuned by the EG length, which provides useful design guidelines for the development of effective NF-based vaccines.

**Keywords:** peptide; nanofibers; poly(ethylene glycol); antigen delivery; immune stimulation
