Background and Objective: Gestational diabetes is one of the most common metabolic disorders during pregnancy. In order to find out a simple and cost effective method with acceptable sensitivity and specificity, fasting plasma glucose (FPG) and one hour 50-g glucose challenge test (OGCT) were compared in patients with gestational diabetes mellitus (GDM). Materials and Methods: In this prospective cohort study, pregnant women without preexisting diabetes underwent FPG and OGCT tests between 24 and 28 weeks of gestation. If the OGCT threshold values exceeded ≥ 130 mg/dl, the 100g oral glucose tolerance test (OGTT) was performed using Carpenter and Coustan criteria. Receiver operating characteristic (ROC) analysis was used to evaluate the performance of the two tests. Results: GDM was diagnosed in 7.3% and impaired glucose tolerance in 3.2%. The best cut-off points for GCT and FPG were 134mg/dl(sensitivity: 99.24%, specificity: 76.57%) and 87mg/dl(sensitivity: 80.15%, specificity: 85.62%).By using GCT, an optimal cut-off values of GCT<135mg/dl (sensitivity: 96.95%) to rule out GDM and values ≥ 165mg/dl (specifity: 96.10%) to rule in GDM, would eliminate the need for the OGTT in 80.1% women (misclassification rate: 3.83%). By using FPG, an optimal cut-off values of <76mg/dl (sensitivity: 95.42%) to rule out GDM and values ≥ 91mg/dl (specifity: 95.56%) to rule in GDM, would eliminate the need for the OGTT in 51% women (misclassification rate: 4.43%). Conclusion: The results showed that the best test for predicting macrosomia, preterm delivery and caesarian section is OGCT and for preeclampsia and respiratory distress is FPG. As OGCT can decrease the necessity of OGTT performance with lower misclassification rate comparing to FPG, OGCT would be the best screening test for GDM in Iran.