2.3. Synthesis of AT13148 2
2.4. Synthesis of compound 3 (the probe)
To a solution of 2 (116 mg, 0.3 mmol) in dry DCM (15 mL) was added bromotrimethylsilane (0.42 mL, 3.0 mmol) dropwise under nitrogen at 0 °C. The reaction mixture was stirred for 48 h at room temperature. The precipitate was collected by filtration and washed with diethyl ether three times and dried to afford compound 3 as a white solid (yield 90%). 1H NMR (d6-DMSO, 600 MHz, ppm, Figure S5): 2.41 (s, 3H), 7.38 (d, 2H), 7.53 (t, J = 7.2 Hz, 1H), 7.74 (d, 1H), 7.69 (t, J = 6.6 Hz, 1H), 8.03 (d, 2H), 8.11 (d, 1H). MS-ESI (Figure S6): m/z = 329.88 [M−H]−. 31P NMR (δ ppm, in d6-DMSO, Figure S7): −5.39. The synthesis of the probe is somatostatin shown in Scheme 1.
2.5. Alkaline phosphatase (ALP) ELISA Kit assay
Detection of ALP using the ELISA Kit assay was conducted according to the instructions of the ELISA Kit, which involves 11 steps.