Ar2000 tlc scanner

To determine apparent molar activity (AMA), a DOTA titration was performed using ⁴³Sc or ⁴⁷Sc, after which, the half-maximum effective concentration (EC₅₀) for complete complexation was determined by taking the best-fit values of a transform of the log(µmol) versus percent radiolabeled, performed using Prism 8 software. Then, the average activity added was divided by EC₅₀ multiplied by 2, shown in Eq. (3) below. $$AMA=\frac{\text{Average activity}}{\left(E{C}_{50}\times 2\right)}$$
Details of the titration are provided in the supplemental information; section i of the supplemental and Table S6. Analysis of the samples was performed via instant thin-layer chromatography by spotting 1 µL of sample on an iTLC-SG paper and developing in 1 M citrate buffer. An Eckert & Ziegler AR2000 TLC scanner (Berlin, Germany) was used for TLC analysis.

Stock solutions of
the ligands (10^–3 M) were prepared in ultrapure
deionized H₂O and diluted appropriately to give a serial
dilution series (10^–4–10^–6 M). Concentration-dependent radiolabeling was performed by the addition
of [²⁰³Pb]Pb²⁺ (10 μL, 124 kBq) to a solution
containing the ligand (10 μL, 10^–3–10^–6 M) diluted in NH₄OAc buffer (80 μL,
1 M, pH = 7). Water replaced the ligands in the negative control.
All of the radiolabeling for the cyclen-based derivatives was performed
at room temperature and monitored at 5 min and 1 h time points whereas
heating at T = 80 °C was also employed for TACD3S,
TRI4S, and TE4S. All radiolabeling reactions were repeated at least
in triplicate.
Radiochemical incorporation (RCI) was determined
via instant thin-layer chromatography (iTLC) with silicic acid (SA)-impregnated
paper TLC plates (iTLC-SA, Agilent Technologies, USA). Ethylenediamine
tetraacetic acid (EDTA, 50 mM, pH = 5.5) was used as the eluent. Under
these conditions, free [²⁰³Pb]Pb²⁺ migrates
with the solvent front (R_f = 1) while
the [²⁰³Pb]Pb²⁺ complexes remain at the baseline
(R_f = 0). The iTLC plates were analyzed
on an Eckert & Ziegler AR-2000 TLC scanner, and all the data were
processed with Eckert & Ziegler WinScan software. Representative
TLC radiochromatograms are presented in Figure S20.

The radiolabeling yield of [¹³¹I]I-peptide was determined by TLC using Whatmann 3 MM strips and MeOH/H₂O (95:5), as eluent. For [^99mTc]Tc(CO)₃-peptide, TLC-SG (Al) strips (Merck; Burlington, MA, USA) using ACN/H₂O (95:5) as eluent were used; and for only [^99mTc][Tc(CO)₃]⁺ complex the radiolabeling yield was evaluated using TLC-SG (Al) strips and 0.9% NaCl, as eluent. The radioactivity was determined by AR 2000 TLC-Scanner (Eckert & Ziegler; Berlin, Germany).
RP-HPLC analyzes were performed on a 1290 Infinity II UHPLC system (Agilent Technologies; Saint Clair, CA, USA) fitted with a radioactivity detector (Eckert & Ziegler; Berlin, Germany) and Open Lab ECM data system (Agilent Technologies; Saint Clair, CA, USA) to confirm the yield and purity of both radiolabeled peptides. Analyzes were performed using a Phenomenex C18 column (150 × 46 mm; 5 µm particle size; 300 Å pore size), detection at λ = 220 nm, using 0.1% TFA:H₂O as solvent A and 60% acetonitrile/0.1% TFA:H₂O as solvent B. A gradient of 5% to 95% of solvent B in 30 min was used at a flow rate of 1.0 mL·min⁻¹.