The effects of temperature on the key parameters of a prototype GaAs 55Fe radioisotope X-ray microbattery were studied over the temperature range of −20 °C to 70 °C. A p-i-n GaAs structure was used to collect the photons from a 254 Bq 55Fe radioisotope X-ray source. Experimental results showed that the open circuit voltage and the short circuit current decreased with increased temperature. The maximum output power and the conversion efficiency of the device decreased at higher temperatures. For the reported microbattery, the highest maximum output power (1 pW, corresponding to 0.4 μW/Ci) was observed at −20 °C. A conversion efficiency of 9% was measured at −20 °C.