An incubator is a womb-like environment for preterm infants which improve their chances of survival. Monitoring the incubator environment is crucial as it play vital role towards the growth and survival of the preterm infant. This paper presents the design, calibration, implementation and performance evaluation of a real-time neonatal incubator assistant (NIA) using Internet-of-Things (IoT) technology for monitoring the environment of a standard preterm infant incubator. The six parameters under close monitoring are: 1). temperature, 2). humidity, 3). heart beat rate per minute, 4). heat pulse per minute, 5). sound level, and 6). air quality. The proposed IoT-based NIA consist of three main systems, namely: 1). The IoT-based hardware module; 2). A web and database system; and 3). Wi-Fi wireless remote monitoring system with a software application for online monitoring of the preterm infant incubator via a mobile application (Mobile App). The satisfactory validation performance evaluations of the designed and constructed IoT-based NIA shows that it can be deployed in a hospital clinical setting for preterm infant monitoring in an incubator.