Spe cartridge

200 μL of delipidated serum samples [11 (link)] were diluted 5 times in 0.1% TFA prior to C18 solid phase extraction (SPE). The SPE cartridges (PID# WAT020515, Waters) were washed stepwise in 90% ACN and finally equilibrated 3 times in 0.1% TFA before the loading of diluted serum. Then, the SPE cartridge was washed three times with 0.1% TFA to remove binding of non-specific proteins on the C18 reverse phase and elution carried out with 70% ACN, 0.1% TFA.

Standard reference material (SRM) solutions made from inorganic fluorine “fluoride standard solution” (nr. 1.19814.0500) and “anion multielement standard I” (nr. 1.11437.0500) were purchased from Merck KGaA, Darmstadt, Germany. PFOA in solid form was bought from Aldrich, now part of Sigma-Aldrich (St. Lousi, MO, United States). Standard solutions of PFAS were purchased from Wellington Laboratories (Guelph, Canada). Potassium and magnesium salts (KCl, reagent grade; MgCl₂ x 6 H₂O, reagent grade) were purchased from Scharlau (Barcelona, Spain). Sodium chloride (NaCl) was purchased from VWR Chemicals (Radnor, PA, US). Calcium chloride (CaCl₂ x 2 H₂O) was purchased from Merck.
Methanol (MeOH) was purchased from Fisher Scientific (Waltham, MA, United States). Water for sample preparation and the CIC (absorber, rinsing, eluent solutions) was purified with a MilliQ system (18.2 MΩ resistance). Argon and oxygen for the CIC were purchased from AGA (Lidingö, Sweden). Methyl tert-butyl ether (MTBE) and tetrabutyl-ammonium sulfate (TBA), used for IPE, were purchased from Sigma-Aldrich. SPE cartridges (6 mL, 150 mg sorbent, 30-μm particle size) with WAX sorbent were purchased from Waters Corporation, Milford, MA, US. GF/F glass fiber filters (150 mm, 0.7 μm) were purchased from Whatman (Chicago, IL, US).

Unless otherwise stated, lyophilized samples were resuspended in 8 M Urea in 50 mM ammonium bicarbonate, reduced with 5 mM TCEP (Sigma-Aldrich) for 30 min at 37 °C on a thermomixer (Eppendorf) and subsequently alkylated with 10 mM IAA (Sigma) at room temperature in the dark. Urea was first diluted to a concentration of 5.5 M and Lys-C (Wako) at a 1:100 wt/wt ratio was added for 2.5–3 h at 37 °C. Subsequently, urea was diluted to 1 M and samples were further digested by addition of trypsin (Promega) at 1:50 wt/wt ratio overnight at 37 °C. Digested samples were acidified by addition of formic acid and purified using either SPE cartridges (Waters) or microspin columns (The Nest Group). Eluates were dried, resuspended in acetonitrile 2–5% and formic acid 0.1%, typically with addition of iRT peptides (Biognosys), sonicated and centrifuged for 5–10 min at 10,000 g before MS injection.

The cyclotron parameters used in the production of carrier-added [¹⁸F]F₂ are summarized in Supplemental Table 1. The target gas ([¹⁸O]O₂ > 98%) used for the first bombardment on the cyclotron is obtained from Campro Scientific (Berlin, Germany). The synthesis is carried out using a custom-build synthesis platform (see Supplemental Figs. 1 and 2). An overall synthesis flowchart can be found in Supplemental Fig. 3. The precursor 2 for radiolabeling is obtained from ABX advanced biochemical compounds (Radeberg, Germany), standard reagents are obtained from Sigma-Aldrich, sterile solutions from the Aarhus University Hospital Pharmacy (Aarhus, Denmark), SPE cartridges and sterile filters from Waters, and the high-performance liquid chromatography (HPLC) column is obtained from Phenomenex. The radiosynthesis is described in detail in the Supplemental Data. Following completed deprotection of [¹⁸F]3 (see Scheme 1), the crude product is purified by semi-preparative radio-HPLC using a Spherisorb ODS column (250 × 10 mm) and sodium dihydrogen phosphate buffer (70 mm, sterile) as eluent at a flow rate of 3 ml/min. [¹⁸F]1 elutes at 11–13 min and is collected directly into the final sterile product vial via a Millipore-GS 0.22 μm sterilizing filter.

All reagents were commercially acquired and utilized without further purification unless otherwise stated. 4,7,13,16,21,24-Hexaoxa-1,10-diazabicyclo [8.8.8] hexacosane (K222, 98.0 %) was purchased from ABX (Germany). Anhydrous acetonitrile and potassium carbonate (K₂CO₃) were purchased from Sigma (America). All SPE cartridges were purchased from Waters Corporation (USA). To activate Sep-Pak QMA, Sep-Pak C18, and Sep-Pak PS-2, we flushed them with 5 mL absolute ethanol followed by 10 mL water.