*2.3. bi-HNTs Synthesis*

bi-HNTs were synthesized using the same methods as described in a previous study (Figure 2) [16]. Briefly, HNTs (Sigma Aldrich, St. Louis, MO, USA) were reacted at reflux with *N*-[3-(trimethoxysilyl) propyl] ethylenediamine (DAS) (Sigma Aldrich, St. Louis, MO., USA) in toluene for 24 h. Then, the HNT-DAS composite was reacted with FA (Sigma Aldrich, St. Louis, MO, USA) in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) (Sigma Aldrich, St. Louis, MO, USA) in DI water overnight. Finally, the HNT-DAS-FA composite was complexed with FITC (Sigma Aldrich, St. Louis, MO, USA) in acetone (Granger, Lake Forest, IL, USA) overnight.

**Figure 2.** Schematic representation of the conjugation of both FA and FITC to DAS which is attached to the HNT surface. Methotrexate (MTX) was vacuum-doped into the HNT lumen. DAS = *N*-[3- (trimethoxysilyl)propyl) ethylenediamine, FA = folic acidity, FITC = fluorescein isothiocyanate.

#### *2.4. bi-HNTs Characterization*

2.4.1. Fourier-Transform Infrared Spectroscopy (FTIR)

The Infrared spectrum was recorded at a resolution of 4 s−<sup>1</sup> with 16 scans using a Thermo Scientific NICOLET™ IR100 FT-IR Spectrometer (Thermo Fisher Scientific; Waltham, MA, USA). Thermo Scientific OMNIC™ software (Waltham, MA., USA) was used to study the stretched bands.
