2545 pump gradient module

All chemicals, reagents, and solvents for the synthesis were purchased from Sigma Aldrich (St. Louis, MO, USA), Merck (Darmstadt, Germany) and Iris Biotech (Marktredwitz, Germany). IRDye^® 700DX NHS Ester was obtained from LI-COR, Inc. (Lincoln, NE). Analytical and preparative HPLC−MS was carried out on a Waters AutoPurification system (3100 Mass Detector, 2545 Pump Gradient Module, 2767 Sample Manager, and 2998 PDA detector).
The peptidomimetic PSMA binding motif glutamate-urea-lysine binding motif (Glu-NH-CO-NH-Lys-2-naphthyl-L-Ala-cyclohexane) was synthesized by solid-phase peptide chemistry according to previously published methods (23). The PSMA binding motif (1 mg, 1.5 μmol), was conjugated to IRDye700DX-NHS (1.9 mg, 1 μmol) in buffer phosphate 0.1 M, pH = 8 The reaction was stirred for 2 h at room temperature followed by preparative HPLC-MS. The details of the preparation and characterization of IRDye700DX-PSMA are reported in the Supplementary Materials.

Chemicals were
purchased from Sigma Aldrich Co. NMR spectra were
recorded at 310 K on a Bruker AVANCE 600 spectrometer operating at
14 T (corresponding to 600 and 150 MHz ¹H and ¹³C Larmor frequencies, respectively). Analytical and preparative HPLC–MS
was carried out on a Waters Auto Purification system (3100 Mass Detector,
2545 Pump Gradient Module, 2767 Sample Manager, and 2998 PDA detector).
The purity was double checked by analytical HPLC using an Atlantis
C18, 3.5 μm, 4.6 mm × 150 mm column and 0.1% TFA in water
(solvent A) and acetonitrile (solvent B); applying a gradient of CH₃CN in H₂O (0.1%TFA) from 5 to 80% in 15 min and
from 80 to 100% in 5 min, flow 1 mL min^–1 (method
1). pH measurements were made by using an AS pH meter equipped with
a glass electrode.