**2. Results and Discussion**

#### *2.1. Preparation of Bifunctional Chelators NOTA-PODS and NODAGA-PODS*

The aluminum fluoride-18 ([18F]AlF) radiolabeling procedure has generated significant interest by combining the convenient and straightforward metal-based radiochemistry (e.g., one-pot reaction, minimal purification steps, aqueous solution compatibility) and the excellent decay characteristics of fluorine-18 (97% positron emission, low positron energy, half-life of 109.8 min). This method was used to directly radiolabel small molecules, peptides and proteins containing macrocyclic chelators such as NOTA or NODA [16,17]. Accordingly, in this work, the NOTA and NODAGA chelators were selected for the [18F]AlF radiolabeling of affibody molecule ZEGFR:03115. However, instead of the conventional maleimide, thiol-reactive bifunctional chelators for the conjugation to the cysteine-containing small protein were developed by functionalizing the macrocycles with PODS. The reaction between PODS and the NHS ester of either NOTA or NODAGA was performed in DMF in the presence of base (Scheme 1). The products were obtained in a quantitative yield (Figure S1); NOTA-PODS and NODAGA-PODS were used for the subsequent conjugation reaction without further purification. Alternatively, pure bifunctional chelators could be isolated by semi-preparative RP–HPLC using formic acid instead of TFA in the mobile phase (Figures S2–S4). Of note, when NOTA-PODS solutions in DMF were stored at −20 ◦C, noticeable signs of degradation were detected already after 2 months. Conversely, NODAGA-PODS in DMF showed good stability for at least 10 months (Figure S5).

**Scheme 1.** Synthesis of NOTA-PODS and NODAGA-PODS.
