Whatman no 2

Typically, 510 MBq of ¹¹¹InCl₃ solution in 0.2 M HCl in conical vials were dried under a flow of nitrogen and residues were dissolved in 1500 μl of ammonium acetate buffer (pH = 5.5). 500 μl aliquots of ammonium acetate buffer were added to the vials containing lyophilized immunoconjugates (three different DOTA: antibody ratios) and the vials were mixed using pipetting up and down (10-20×) to dissolve immunoconjugates. Then 500 μl (170 MBq) aliquots of ¹¹¹In solution were added to vials and after mixing gently for 5 min by up and down pipetting, mixtures were incubated at 37°C for 1–2 h.
The radiochemical purity of the products was determined by ITLC on Whatman No.2, using 10 mM DTPA as mobile phase and HPLC on a C-18 column using a gradient system as a reported routine method in this laboratory
[27 ]. To increase the radiochemical purity, the radioimmunoconjugates were purified by chromatography using PD-10 columns (GE healthcare) and ammonium acetate buffer (pH = 7.0) as eluting solvent. The final solutions were then passed through 0.22 micron biological filters for stability and biodistribution studies. Radiolabeling of the immunoconjugates with ⁹⁰Y and quality control of the resulting radioimmunoconjugate were carried out by the same method which was described for the preparation and quality control of ¹¹¹In-DOTA-rituximab.

Fresh P. densiflora and P. koraiensis needles were rinsed with distilled water. After removing moisture, the needles were dried in a heat dry machine (KED-066A, C&T Co., Gwangju, Korea) at 30 °C for seven days or 90 °C for three days without light. Then, dried samples were coarsely ground using a commercial grinder and passed through a 100-mesh sieve. Subsequently, the resulting powder (10 g) from each sample was dissolved in 200 mL of 80% aqueous methanol (v/v) and agitated using a shaker (Daewonsci Inc., Bucheon, Korea) at 20 °C for 24 h. Next, the solution was filtered through qualitative filter papers (Whatman No. 2, Maidstone, UK). Finally, the solvent was removed using a vacuum rotary evaporator (Eyela, Tokyo, Japan). To remove residual solvent, the extracts were dried using a freeze dryer (IlShinBioBase Inc., Dongducheon, Korea) and maintained at 15 °C until use for non-volatile assay.

Before the extraction of phenolic compounds, each lyophilized leaf sample was powered and sieved using a 900 μm sieve. The extraction was performed as previously described by Vinha et al. (2002) (link). Briefly, about 1.5 g of the powdered leaf samples were weighed in quadruplicates. Each sample was separately mixed with 50 mL of methanol (99.96%, Aldrich) at 150 rpm for 1 h (room temperature). The obtained methanolic extracts were filtered through a Whatman No.4 paper and evaporated (Stuart RE3000, UK) to dryness under reduced pressure (35°C). After dissolution in 2 ml methanol (99.96%, Aldrich) and filtration (Whatman No. 2), an aliquot of 20 μl of the obtained extracts was analyzed by HPLC.