Pafolacianine

Guide (The following is our recommended protocol. This protocol is provided as a guide only and should be modified to suit your specific needs)^[3].
1. Preparation of dye stock solution
1) Pafolacianine is stored at 1508 μM in a -80°C freezer.
2. Preparation of dye working solution
1) Dilute Pafolacianine in 5% glucose solution within 24 h before use.
2) For the in vivo mouse xenograft model and the ex vivo porcine lung model, prepare working solutions of 0.05 mg/kg, 0.025 mg/kg, and 0.0125 mg/kg by diluting the stock solution in 5% glucose solution.
3. Specific staining steps
3.1 In vivo mouse xenograft model
1) 100 μL of cell mixture (70% KB cell suspension containing 1.0×10⁶ cells in phosphate buffered saline, 30% Matrigel) is subcutaneously inoculated into the right thigh of 8-12 week old female athymic nude mice to establish a subcutaneous xenograft tumor model.
2) Mice are intravenously injected with 0.05 mg/kg, 0.025 mg/kg, or 0.0125 mg/kg of Pafolacianine, or injected with 5% glucose solution as a negative control.
3) Intermittent imaging is performed using UCF at 5, 10, and 15 mm above the tumor (right thigh) and normal limb (left thigh, background) at 6, 24, 48, and 72 h after injection, respectively.
4) Additional NIR imaging is performed using an in vivo whole mouse fluorescence imaging system (Maestro) with 710-760 nm excitation and >800 nm detection.
5) To evaluate biodistribution, mice are sacrificed 24 h after injection, tumors and organs are excised, and fluorescence intensity is assessed using the Maestro system and UCF.
3.2 Ex vivo peribronchial tumor pig model
1) Mice bearing KB tumors with a diameter >10 mm are injected intravenously with 0.025 mg/kg or 0.05 mg/kg of Pafolacianine 24 h before tumor resection.
2) Pafolacianine-loaded KB tumors are excised and transplanted into the peribronchial sites (carina and peripheral airways) of ex vivo porcine lungs. A bronchoscope (BF-P190) is inserted through the endotracheal tube, and the UCF is inserted into the working channel of the bronchoscope.
3) Increase the laser power to 75 mW, acquire NIR fluorescence images in a dark room, and confirm the absence of bronchial wall autofluorescence by UCF using the same settings before tumor placement.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.