Ptfe syringe filter

Poly(ethylene glycol) diacrylate (M_n 700) (PEG₇₀₀DA), 2-aminoethyl methacrylate hydrochloride (AEM), and diphenyl (2,4,6-trimethylbenzoyl)-phosphine oxide (TPO) were purchased from Sigma-Aldrich. Thermo Scientific Dylight 488, PTFE syringe filters (13 mm membrane, 0.22 μm pore size), sterile water, and methanol were obtained from Fisher Scientific. Conventional filters (2 μm) were purchased from Agilent, polyvinyl alcohol (Mw 2000) (PVOH) was purchased from Acros Organics, and Luvitec (MW 64 kDa) was purchased from BASF. PRINT molds (80 nm x 320 nm, 80 nm x 5000 nm, and 200 nm x 200 nm) were obtained from Liquidia Technologies. Tetraethylene glycol monoacrylate (HP₄A) was synthesized in-house as previously described [39 ].

Poly(ethylene glycol) diacrylate (Mn 700) (PEG700DA), 2-aminoethyl methacrylate hydrochloride (AEM), and diphenyl (2,4,6-trimethylbenzoyl)-phosphine oxide (TPO) were purchased from Sigma-Aldrich. Thermo Scientific Dylight 488, PTFE syringe filters (13 mm membrane, 0.22 μm pore size), sterile water, and methanol were obtained from Fisher Scientific. Conventional filters (2 μm) were purchased from Agilent, polyvinyl alcohol (Mw 2000) (PVOH) was purchased from Acros Organics, and Luvitec (MW 64 kDa) was purchased from BASF. PRINT molds (80 nm x 320 nm, 80 nm x 5000 nm, and 200 nm x 200 nm) were obtained from Liquidia Technologies. Tetraethylene glycol monoacrylate (HP4A) was synthesized in-house as previously described [30 ].

We synthesized FITC labeled PAH using the previously reported method^{56 (link),57 (link)}. Briefly, we dissolve 20 mg of FITC in 2.5 mL of dimethyl sulfoxide (DMSO). In a separate vial, 2.5 g of PAH (M.W. ~17500 Da) was dissolved in 30 mL of 18.2 MΩ-cm deionized (DI) water and adjusted to pH of 9 using 6 M NaOH. Then, FITC in DMSO solution is carefully poured into PAH solution and stirred for 2 days in the dark at room temperature. The resulting solution was subsequently dialyzed to DI water with 7 kDa MW cut-off tubing (SnakeSkin, ThermoFisher) for 7 days. After dialysis, solution was filtered with 0.2 µm PTFE syringe filter (ThermoFisher) prior to usage.

A. flavus and F. proliferatum isolates were cultured on rice medium (2% agar-VWR, BDH chemicals; 2% rice powder) at 30 °C/10 days. Five grams of rice medium (using cork borer 4 mm to collect medium) containing fungi were added to previously weighed 15 mL tubes. Aflatoxin B1 and Fumonisin B1 were extracted with 2.5 mL of chloroform (HPLC grade, VWR International, Leuven, Belgium), vortexed (VM-1000) for 1 min, shaken 200 rpm/60 min (S100H, Elmasonic, AHRD), and then centrifuged at 3000 rmp/5 min (EBA200). The supernatant was transferred to a 5 mL glass tube and dried gently under 40 °C by nitrogen (MRC, AHRD). One mL mobile phase MeOH: H₂O (0.1% acid formic) 7/3: v/v was added to such dried glass tube and vortexed. This solution was filtered by syringe filters 0.45 μm (PTFE syringe filter, Thermo Fisher) and transferred to a new vial.

The extract (10 mg) was subjected to solid phase extraction (SPE) using a Hypersep C18 cartridge (Thermo Scientific, Waltham, MA, USA) eluting sequentially with 70, 80, 90, and 100% methanol (MeOH). Fraction 1 (1.5 mg), fraction 2 (1.8 mg), fraction 3 (2.1 mg), and fraction 4 (4.2 mg) were collected, and stock solutions (0.1 mg/mL) were prepared for further analysis. The sample solution and blank control were all stored at 4 °C and filtered through a 0.22 μm PTFE syringe filter (Thermo Scientific, Waltham, MA, USA) prior to analysis by HPLC-DAD-MS. Analysis was carried out using an Agilent HPLC 1260 binary pump and a Sunfire C18 column (2.1 × 150 mm, 3.5 µm) (Waters, Milford, MA, USA). The diode array detector (DAD) was set at λ = 210, 254, 269, and 310 nm. The known, or suspected, major adducts for all samples were registered in the positive electrospray ionization (ESI) mode either as [M+H]⁺ or [M+Na]⁺. The mobile phase used was a gradient of 1% formic acid in water and acetonitrile (solvents A and B, respectively) at a flow rate of 0.2 mL/min. Elution was performed as follows; Fraction 1 (0 to 30 min—60 to 80% solvent B), fraction 2 (0 to 30 min—70 to 90% solvent B), fraction 3 (0 to 30 min—80 to 100% solvent B), and fraction 4 (0 to 30 min—100% solvent B).