Vaccine development for Plasmodium vivax malaria is underway. A model to assess the protective efficacy of vaccine candidates in humans is urgently needed. Given the lack of continuous P. vivax cultures, we developed a system to infect Anopheles albimanus mosquitoes using blood from P. vivax-infected patients and determined parameters for challenge of malaria-naive volunteers by mosquito bite. Absence of co-infections in parasitized blood was confirmed by tests consistent with blood bank screening. A total of 119 experiments were conducted using batches of 900–4,500 mosquitoes fed by an artificial membrane feeding method. Optimal conditions for mosquito probing and infection were determined. Presence of oocyst and sporozoites were assessed on Days 7–8 and 14–15, respectively, and conditions to choose batches of infected mosquitoes for sporozoite challenge were established. Procedures to infect volunteers took a 2-hour period including verification of inoculum dose. Anopheles albimanus mosquitoes represent a valuable resource for P. vivax sporozoite challenge of volunteers.