Kinase experiment:

Rhodamine 800 (Rh800) and indocyanine green (ICG) are dissolved in PBS and their concentrations are set to 40 nM. To 3 mL of Rhodamine 800 solution (40 nM/PBS), 100 μL of p-sulfonatocalix[n]arenes (S[n]) (1 mM) is added. To this solution, microliter aliquots of acetylcholine (ACh) (350 mM) are added under stirring. After 2 min stirring, fluorescence spectra are measured. For the competitive fluorophore displacement by other neurotransmitters, amino acids, ammonium chloride, and choline, similar procedure as the above method is applied[2].

Rhodamine 800 is a near-infrared fluorescent dye.

Rhodamine 800 has different optical absorption spectra in various organic solvents and in aqueous solution in the visible and near-infrared regions. The absorption maximum at 683 nm in acetone shifts to 696 nm in DMSO without a significant change of the overall spectral profile. The fluorescence intensity of Rhodamine 800 varies markedly with the solvent, and decreases to less than one-fifth when the solvent is changed from acetone to water without a significant change of the emission peak[1].

[1]. Sakanoue J, et al. Rhodamine 800 as a probe of energization of cells and tissues in the near-infrared region: a study with isolated rat liver mitochondria and hepatocytes. J Biochem. 1997 Jan;121(1):29-37. [2]. Jin T. et al. Near-infrared fluorescence detection of acetylcholine in aqueous solution using a complex ofrhodamine 800 and p-sulfonatocalix[8]arene. Sensors (Basel). 2010;10(3):2438-49.